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Review Paper

Developments in industrially important thermostable enzymes:a review

G.D. Haki, S.K. Rakshit *

Bioprocess Technology Program, Asian Institute of Technology (AIT), P.O. Box 4, Klong Luang, Pathumthani 12120, Thailand

Received 18 November 2002; received in revised form 3 January 2003; accepted 13 January 2003

Abstract

Cellular components of thermophilic organisms (enzymes, proteins and nucleic acids) are also thermostable. Apart from high

temperature they are also known to withstand denaturants of extremly acidic and alkaline conditions. Thermostable enzymes arehighly specific and thus have considerable potential for many industrial applications. The use of such enzymes in maximising re-

actions accomplished in the food and paper industry, detergents, drugs, toxic wastes removal and drilling for oil is being studied

extensively. The enzymes can be produced from the thermophiles through either optimised fermentation of the microorganisms or

cloning of fast-growing mesophiles by recombinant DNA technology. In this review, the source microorganisms and properties of

thermostable starch hydrolysing amylases, xylanases, cellulases, chitinases, proteases, lipases and DNA polymerases are discussed.

The industrial needs for such specific thermostable enzyme and improvements required to maximize their application in the future

are also suggested.

Ó 2003 Elsevier Science Ltd. All rights reserved.

Keywords: Thermostable enzymes; Thermophilic microorganisms

1. Introduction

The role of enzymes in many processes has been

known for a long time. Their existence was associated

with the history of ancient Greece where they were using

enzymes from microorganisms in baking, brewing, al-

cohol production, cheese making etc. With better

knowledge and purification of enzymes the number of

applications has increased manyfold, and with the

availability of thermostable enzymes a number of new

possibilities for industrial processes have emerged.

Thermostable enzymes, which have been isolated mainly

from thermophilic organisms, have found a number of commercial applications because of their overall inher-

ent stability (Demirijan et al., 2001). Advances in this

area have been possible with the isolation of a large

number of beneficial thermophilic microorganisms from

different exotic ecological zones of the earth and the

subsequent extraction of useful enzymes from them

(Burrows, 1973; Antranikian et al., 1987; Groboillot,

1994; Bharat and Hoondal, 1998; Bauer et al., 1999;

Kohilu et al., 2001).

While the most widely used thermostable enzymes are

the amylases in the starch industry (Poonam and Dalel,

1995; Crab and Mitchinson, 1997; Emmanuel et al.,

2000; Sarikaya et al., 2000), a number of other appli-

cations are in various stages of development. In the food

related industry, they have been used in the synthesis of

amino acids (Satosi et al., 2001). In the petroleum,

chemical and pulp and paper industries, for example,

thermostable enzymes have been used for the elimina-

tion of sulphur containing pollutants through the

biodegradation of compounds like dibenzothiophene(Bahrami et al., 2001), in the production of 1,3-pro-

panediol from glycerol and in replacing polluting

chemical reagents causing toxic products (Peter et al.,

2001). Currently, a number of publications have exten-

sively discussed developments in this area. Adaptation

of extremophiles to hot environments (Danson et al.,

1992; Stetter, 1999), production of heat-stable enzymes

from thermophiles and hyperthermophiles (Knor, 1987;

Jakob, 1989; Huber and Stetter, 1998; Niehaus et al.,

1999), structure and function relationships of thermo-

zymes (heat-tolerant enzymes) (Zeikus et al., 1998a,b)

*Corresponding author. Tel.: +66-2-524-6115; fax: +66-2-524-6111.

E-mail address: [email protected] (S.K. Rakshit).

0960-8524/03/$ - see front matter Ó 2003 Elsevier Science Ltd. All rights reserved.

doi:10.1016/S0960-8524(03)00033-6

Bioresource Technology 89 (2003) 17–34

http://mail%20to:%[email protected]/





and biotechnological and industrial applications of

thermostable enzymes (Franks, 1993; Lasa and Beren-

guer, 1993; Leuschner and Antranikan, 1995; Cowan,

1996; Holst et al., 1997; Hough and Danson, 1999;

Eichler, 2001) are among the topics that have been

studied.

In the present review, an attempt is made to docu-

ment the research activities conducted in the area and

indicate the source microorganisms of some important

thermostable enzymes. The need for thermostable cata-

lysts, the optimum conditions for an efficient catalytic

activity of the enzymes and current industrial applica-

tions are presented. Besides discussing the reason for the

thermostable character of the enzymes, possible im-

provements and a number of expected developments are

also suggested in the article.

2. Geothermal sites as sources of extremophiles

In situ temperatures between 80 and 115 °C were

found to be conducive biotopes for a number of hy-

perthermophiles (Huber and Stetter, 1998). Some ex-

amples are mentioned in Table 1. Extremophiles are also

adapted to live at low temperatures in the cold polar

regions, at high pressure in the deep sea and at a very

low and high pH values (pH 0–3 or 10–12), or at a very

high (5–30%) salt concentration (Herbert and Sharp,

1992).

In the last decade, a number of hyperthermophilic

archaea, the least understood domain of life (Woese

et al., 1990; Rotschild and Manicinelli, 2001) have beenisolated and are able to grow around the boiling point of

water (Niehaus et al., 1999). The organisms with the

highest growth temperatures (103–110 °C) are members

of the genera Pyrobaculum, Pyrodictium, Pyrococcus

and Methanopyrus. Within the bacteria, Thermotoga

maritima and Aquifex pyrophilus exhibit the highest

growth temperatures of 90 and 95 °C respectively

(Herbert and Sharp, 1992). These properties imply ex-

tremely important industrial and biotechnological im-

plications due to the fact that enzymes from such

microorganisms can be employed for use in harsh in-

dustrial conditions where their specific catalytic activity

is retained.

3. Resistance of thermophiles to high temperatures and

denaturation

Microorganisms, like all living things, adapt to the

condition in which they have to live and survive. Ther-

mophiles are reported to contain proteins which are

thermostable and resist denaturation and proteolysis

(Kumar and Nussinov, 2001). Specialized proteinsknown as ÔchaperoninsÕ are produced by these organ-

isms, which help, after their denaturation to refold the

proteins to their native form and restore their functions

(Everly and Alberto, 2000). The cell membrane of

thermophiles is made up of saturated fatty acids. The

fatty acid provides a hydrophobic environment for the

cell and keeps the cell rigid enough to live at elevated

temperatures (Herbert and Sharp, 1992). The archae,

which compose most of the hyperthermophiles, have

lipids linked with ether on the cell wall. This layer is

much more heat resistant than a membrane formed of

fatty acids (De Rosa et al., 1994).

The DNA of thermophiles contains a reverse DNAgyrase which produces positive super coils in the

DNA (Lopez, 1999). This raises the melting point of

the DNA (the temperature at which the strands of the

double helix separate) to at least as high as the organ-

Table 1

Examples of sites serving as sources of microorganisms which can provide thermo tolerant enzymes

Source Microorganism Enzyme References

Hot spring Thermus sp. a-Amylase Shaw et al. (1995)

Hot spring Bacillus sp. WN.11 a-Amylase Mamo and Gessese (1999)

Deep sea hydrothermal vent Staphilothermus marinus a-Amylase Canganella et al. (1994)

Marine solfatare Thermococcus litoralis Pullulanase Brown and Kelly (1993)Decomposed plant samples

from a lake

Clostridium absonum CFR-702 Cellulase free xylanase Swaroopa and Krishna (2000)

Hot spring Bacillus thermoleovocans ID-1 Lipase Dong-Woo et al. (1999)

Compost of fermenting citrus

peels, coffee and tea extract

residues

Bacillus strain MH-1 Endochitinase Kenji et al. (1998)

Compost Bacillus stearothermophilus CH-4 b- N -acetylhexosaminidase Kenji et al. (1994)

Korean salt fermented anchovy Bacillus sp. KYJ963 b-Amylase Young et al. (2001)

Deep sea hydrothermal vent Pyrococcus abyssi Alkaline phosphatase Sebastien et al. (2001)

Sediments of hot springs Bacillus sp. 3183 a-Amylase-like pullulanase Badal et al. (1989)

Garbage dump Bacillus circulans Xylanase Ashita et al. (2000)

Compost treated with artichoke

juice

Bacillus sp. Inulinase Jean-Jacques et al. (1987)

18 G.D. Haki, S.K. Rakshit / Bioresource Technology 89 (2003) 17–34



ismsÕ maximum temperature for growth. Thermophiles

also tolerate high temperature by using increased inter-

actions that non-thermotolerant organisms use, namely,

electrostatic, disulphide bridge and hydrophobic inter-

actions (Kumar and Nussinov, 2001).

Thermostable enzymes are stable and active at tem-

peratures which are even higher than the optimum

temperatures for the growth of the microorganisms(Saboto et al., 1999). These authors indicated that rel-

atively few studies have been carried out in this regard

and the only available information is that thermophilic

enzymes are more rigid proteins than their mesophilic

counterparts. A clearer understanding of this capacity

should be possible with new methodologies that clearly

indicate changes in protein structure.

The effect of structural fluctuations of a-amylases

of mesophilic and thermophilic sources were studied

and reported by Fitter et al. (2001). While determining

the conformational entropy of both enzymes, the folded

state showed a higher structural flexibility for the

thermophilic protein than the mesophilic homologue. It

has, thus, been assumed that a mechanism characterized

by entropic stabilization could be responsible for the

higher thermostability of the thermophilic enzyme.

4. General advantages of enzymes from thermophiles

Thermostable enzymes are gaining wide industrial

and biotechnological interest due to the fact that their

enzymes are better suited for harsh industrial processes

(Leuschner and Antranikan, 1995; Fredrich and

Antrakian, 1996; Diane et al., 1997; Zeikus et al.,

1998a,b). Some biocatalytic conversions and industrial

applications of thermostable enzymes are presented in

Table 2.

One extremely valuable advantage of conducting

biotechnological processes at elevated temperatures is

reducing the risk of contamination by common meso-philes. Allowing a higher operation temperature has also

a significant influence on the bioavailability and solu-

bility of organic compounds and thereby provides effi-

cient bioremediation (Becker, 1997). Other values of

elevated process temperatures include higher reaction

rates due to a decrease in viscosity and an increase in

diffusion coefficient of substrates and higher process

yield due to increased solubility of substrates and

products and favorable equilibrium displacement in

endothermic reactions (Mozhaev, 1993; Krahe et al.,

1996; Kumar and Swati, 2001). Such enzymes can also

be used as models for the understanding of thermosta-

bility and thermo-activity, which is useful for protein

engineering. The following sections describe specific

thermostable enzymes, their sources, characteristics and

application requirements of high temperature resistance.

5. Amylolytic enzymes

The starch industry is one of the largest users of

enzymes for the hydrolysis and modification of this use-

ful raw material. The starch polymer, like other such

Table 2

Bioconversion reactions and applications of thermostable enzymes

Enzyme Temperature range (°C) Bioconversions Applications

a-Amylase (bacterial) 90–100 Starch!dext rose syrups Starch hydrolysis, brewi ng,

baking, detergents

a-Amylase (fungal) 50–60 Starch!dextrose syrups Production of maltose

Pullulanase 50–60 Starch!dext rose syrups Producti on of gluc ose syrups

Xylanase 45–65, 105a

Craft pulp!xylan + lignin Pulp and paper industryChitinase 65–75b Chitin! chitobiose Food, cosmetics, pharmaceuticals,

agrochemicals

Chitin! N -acetyl glucosamine

(chitibiase)

N -acetyl glucosamine!

glucosamine (deacetylation)

Chitin! chitosan (deacetylase)

Cellulase 45–55, 95c Cellulose!glucose Cellulose hydrolysis, polymer

degradation in detergents

Protease 65–85 Protein!amino acids and

peptides

Baking, brewing, detergents, leather

industry

Lipase 30–70 Fat removal, hydrolysis,

interesterification, alcholysis,

aminolysis

Dairy, oleo chemical, detergent,

pulp, pharmaceuticals, cosmetics

and leather industry

DNA polymerase 90–95 DNA amplification Genetic engineering/PCRaXylanase from Thermotoga sp.b Within this range enzyme activity was high.c Cellulases from Thermotoga sp.

G.D. Haki, S.K. Rakshit / Bioresource Technology 89 (2003) 17–34 19



polymers, requires a combination of enzymes for its

complete hydrolysis. These include a-amylases, gluco-

amylases or b-amylases and isoamylases or pullulanases

(Poonam and Dalel, 1995). The enzymes are classified

into endo-acting and exo-acting enzymes. a-amylase is an

endo-acting enzyme and hydrolyses linkages in a random

fashion and leads to the formation of linear and branched

oligosaccharides, while the rest are exo-acting enzymes

and attack the substrate from the non-reducing end,

producing oligo and/or monosaccharides. The starch

hydrolytic enzymes comprise 30% of the world Õs enzyme

consumption (Van der Maarel et al., 2002).

The enzymatic conversion of all starch includes gel-

atinization, which involves the dissolution of starch

granules thereby forming a viscous suspension, lique-

faction, which involves partial hydrolysis and loss in

viscosity, and saccharification, involving the production

of glucose and maltose via further hydrolysis. Gelatini-

sation is achieved by heating starch with water, and

starch is water-soluble only at high temperatures whichare dependent on the source (Rakshit, 1998). For hy-

drolysis of the starch to proceed immediately after gel-

atinization, hence, among other things avoiding a lot of

cooling time, the enzyme has to be thermostable.

A number of attempts were made to isolate and

characterize amylolytic enzymes from diversified sources

(Daniel, 1979; Norman, 1979; Fogarty and Kelly, 1995;

Medda and Chandra, 1980; Morgan and Priest, 1981;

Fogarty, 1983; Krishnan and Chandra, 1983; Rolfsmeir

and Blum, 1995; Piller et al., 1996; Crab and Mitchin-

son, 1997; Rama et al., 1998; Jin et al., 2001; Min Ha

et al., 2001; Stamford et al., 2001; Wojciechowski et al.,

2001; Andersson et al., 2002; Carlsen et al., 2002; Dij-

khuizen et al., 2002; Miura et al., 2002) to meet the

requirements of the starch industry. Table 3 shows

thermostable microbial starch hydrolyzing enzymes and

their properties. Thermostable a-amylases were isolated

a long time ago from Bacillus subtilis, Bacillus amylo-

liquefaciens and Bacillus licheniformis (Underkofler,

1976). The commercially available b-amylases with a

catalytic activity up to 60 °C were isolated from various

Bacillus species (Brown, 1987). They can be used inconjunction with debranching enzymes to produce pure

maltose syrups.

Table 3

Source microorganisms and properties of thermostable starch hydrolyzing enzymes

Enzymes Organism Enzyme properties References

Optimal temperature (°C) Optimal pH

a-Amylase Bacillus amyloliquefaciens 70 7.0 Underkofler (1976)

Bacillus licheniformis 100 6.0–6.5 Viara et al. (1993)

Bacillus stearothermophilus 70–80 5.0–6.0 Vihinen and Mantsala (1990)

Bacillus stearothermophilus 70 – Jeayoung et al. (1989)Bacillus subtilis 70 7.0 Canganella et al. (1994)

Lactobacillus manihotivorans 55 5.5 Aguilar et al. (2000)

Myceliophthora thermophila 100 5.6 Ramkrishna et al. (1993)

Pyrococcus furiosus 100 5.5 Laderman et al. (1993a,b)

Pyrococcus woesei 100 6.5–7.5 Koch et al. (1991)

Staphylothermus marinus 65 5.0 Canganella et al. (1994)

Sulfolobus solfataricus – – Haseltine et al. (1996)

Thermococcus aggreganes 100 5.5 Canganella et al. (1994)

Thermococcus celer 90 5.5 Canganella et al. (1994)

Thermococcus fumicolans 95 4.0–6.3 Estelle et al. (1997)

Thermococcus hydrothermalis 85 4.8–7.8 Estelle et al. (1997)

Thermomyces lanuginosus 60 5.6 Jensen and Olsen (1991)

Thermococcus profoundus 80 4.0–5.0 Kwak et al. (1998)

b-Amylase Bacillus circulans 60 – Poonam and Dalel (1995)

Bacillus cereus var. mycoides 50 – Takasaki (1976a,b)

Bacillus sp. 50 7.5 Young et al. (2001)

Clostridium thermosulphurogenes 75 5.5 Shen et al. (1988)

Clostridium thermosulfurogenes 60 5.8–6.0 Nipkow et al. (1989)

Pullulanase Bacillus sp. 60 – Takasaki (1976a,b)

Pyrococcus furiosus 98 5.5 Brown and Kelly (1993)

Pyrococcus woesi 100 5.5–6.0 Rudiger et al. (1995)

Thermococcus aggregans 100 6.5 Canganella et al. (1994)

Thermus caldophilus GK24 75 5.5 Kim et al. (1996)

Thermococcus celer 90 5.5 Canganella et al. (1994)

Thermococcus hydrothermalis 95 5.5 Gantlet and Duchiron (1998)

Thermococcus litoralis 98 5.5 Brown and Kelly (1993)

Thermotoga maritima MSB8 90 6.0 Bibel et al. (1998)




The pullulanases that were produced in earlier work

on different microorganisms were not very suitable for

operation under the conditions prevailing in the industry

(Nakamura et al., 1975; Konishi et al., 1979; De Mot

et al., 1984a,b; Takizawa and Muroorka, 1985). How-

ever, many thermophilic microorganisms have since

been found to produce pullulanases (Suzuki and Chis-

horo, 1983; Jensen and Norman, 1984; Plant et al., 1986;

Antranikian et al., 1987; Koch et al., 1987; Saha et al.,

1988; Canganella et al., 1994; Swamy and Seenayya,

1996) and b-amylases (Shen et al., 1988; Nipkow et al.,

1989; Swampy et al., 1994; Rina and Geeta, 1996; Rama

et al., 1999; Erra-Pujada et al., 2001).

Termamyl and Fungamyl are two well-known amy-

lolyic enzymes which are now available commercially.

These enzymes are used world-wide for the production

of glucose syrups and syrups with different level of

dextrose equivalent. Hetero-oligosaccharides have been

synthesized by Termamyl obtained from B. licheniformis

(Chitradon et al., 2000). In the presence of soluble starchand added non-starch sugars, oligosaccharides were

produced by the reversed catalytic reaction.

One of the concerns of the starch industry is the

calcium requirement of the a-amylase enzymes and the

formation of calcium oxalate, a substance that may

block process pipes and heat exchangers. Besides, such

accumulation in some products like beer is not accept-

able. Its precipitation can be reduced through a decrease

in calcium ion requirement of enzymes and lowering of

the pH of the production process. Activities of a number

of the starch hydrolyzing enzymes, however, are calcium

dependent although the degree of utilization varies

(Table 4). Production of calcium independent and acid

stable a-amylases has been attempted by protein engi-

neering and as a result Termamyl LCe, which was

produced via site-directed mutagenesis, has shown high

calcium independence (Hashida and Bisgaard-Frantzen,

2000).

Although, raw starch dominates in nature there are

few enzymes that can catalyze its hydrolysis efficiently

(Hamilton et al., 1999). Saccharification of liquefied

starch is carried out at low pH values. However, cur-

rently used thermostable a-amylases are not stable at

such low pH (Crab and Mitchinson, 1997). An eco-

nomical process could be attained through the use of

amylases stable at the saccharification stage. In addition

to this, a one-step process of starch hydrolysis using

amylolytic enzymes would decrease the costs of glucose

production. The anticipated diversity of species of

thermophiles within high temperature environments

(Huber and Stetter, 1998) and the requirements of the

new and improved technological operations for enzymes

with novel and fitting characteristics (Emmanuel et al.,

2000) have placed a challenge both for the scientific and

business community.

6. Thermostable xylanases

Xylan, which is the dominating component of hemi-

celluloses, is one of the most abundant organic sub-

stances on earth. It has a great application in the pulpand paper industry (Dekker and Linder, 1979; Chen

et al., 1997; Lee et al., 1998). The wood used for the

production of the pulp is treated at high temperature

and basic pH, which implies that the enzymatic proce-

dures require proteins exhibiting a high thermostability

and activity in a broad pH range (Jacques et al., 2000).

Treatment with xylanase at elevated temperatures dis-

rupts the cell wall structure. This, as a result, facilitates

lignin removal in the various stages of bleaching. Xyl-

anases for such a purpose (1) must lack cellulytic ac-

tivity to avoid hydrolysis of the cellulose fibers, (2) need

to be of low molecular mass to facilitate their diffusion

in the pulp fibers, and (3) most importantly, high yields

of enzyme must be obtained at a very low cost (Niehaus

et al., 1999). According to these authors all commer-

cially available xylanases can only partially fulfill these

requirements, and the optimum temperature for the

activity of most xylanases is reported to be 50–60 °C

with a half-life of about 1 h at 55 °C (Jacques et al.,

2000). However, some xylanases have been reported to

exhibit higher thermal stability and optimal activity

ranging from 80 to 100 °C (Saul et al., 1995; Zverlov

et al., 1996; Morris et al., 1998).

Table 4

Calcium requirement of industrially important starch degrading enzyme

Enzyme Microorganism Application

temperature range (°C)

Application

pH range

Minimum Ca2þ

dosage (ppm)

Bacterial mesophilic

a-amylase

Bacillus subtilis 80–85 6.0–7.0 150

Bacterial thermophilic

a-amylase

Bacillus licheniformis 95–105 6.0–7.0 20

Fungal a-amylase Aspergillus oryzae 55–70 4.0–5.0 50

Amyloglucosidase Aspergillus niger 55–65 3.5–5.0 0

Pullulanase Bacillus acidopolluliticus 55–65 3.5–5.0 0

Source: Hans (1995).




These enzymes have offered a major step in the re-

duction of chlorine consumption in the bleaching pro-

cess of kraft pulp, thus, lowering environmental

pollution by organic halogens (Vikari et al., 1994; Eriks-

son and Heitmann, 1998). The enzymes are generally

classified in to two families. Family 10 groups (EXs 10)

having high molecular weights (>30 Kda) and Family

11 (EXs 11) which are low molecular weight (<30 Kda)

xylanases (Henrissat and Bairoch, 1993; Dominguez

et al., 1995). Among the two families, the EX 10-fold is

found to be more thermostable (Fontes et al., 1995).

Due to their capacity to enhance the bleaching of kraft

paper, however, EXs 11 have remained more attractive

(Clarke et al., 1997; Morris et al., 1998; Georis et al.,

2000).

Thermostable xylanase were isolated from a numberof bacterial and fungal sources (Table 5). Members of

the Bacillus sp., Streptomyces sp., Thermoascus auran-

tiacus and Fusarium proliferatum have been reported to

produce xylanases which are active at temperatures be-

tween 50 and 80 °C. While the Dictyoglomus sp. were

described to produce xylanases operating at an optimum

temperature of 90 °C, a number of Thermotogales sp.

were reported to secrete thermostable xylanases which

can function at higher temperatures. Alkaliphilic and

cellulase-free xylanases with an optimum temperature of

65 °C from Thermoactinomyces thalophilus subgroup C

(Kohilu et al., 2001) and cellulase-free xylanases

from Clostridium abusonum CFR-702 (Swaroopa and

Krishna, 2000) were also reported recently. From one

Thermotoga sp. an enzyme active at 115 °C was pro-

duced and characterized (Simpson et al., 1991). It was

also possible to produce xylanases in solid-state fer-

mentation, where a decreased cultivation time and an

increased concentration of basal medium improved its

production. This is a promising aspect from an eco-

nomic point of view (Park et al., 2002).

The pulp and paper technology is one of the fastest

growing industries and the use of thermostable xylan-

ases seems attractive since they provide global environ-

mental benefits. However, scaling up of the enzyme

production from the respective microorganisms to the

level required by the industry remains to be seen. It isalso worth mentioning that, extreme thermophiles that

are able to secrete xylanase are few. The search for a

thermophile with high yield of enzyme and the desired

characteristics is still being pursued.

7. Thermostable cellulases

Cellulose, the most abundant organic source of feed,

fuel and chemicals (Spano et al., 1975) consists of glu-

cose units linked by b-1,4-glycosidic bonds in a linear

Table 5

Source microorganisms and properties of thermostable xylanases

Organism Enzyme properties References


Baccillus amyloliquefaciens 80 6.8–7.0 Breccia et al. (1997)

Bacillus circulans 80 6.0–7.0 Dhillon and Khanna (2000)

Bacillus sp. 60–75 8.0–9.0 Gessesse (1998)

Bacillus sp. strain SPS-0 75 6.0 Bataillon et al. (2000)Bacillus subtilis 50 6.0 Paula et al. (2002)

Clostridium abosum 75 8.5 Swaroopa and Krishna (2000)

Dictyoglomus sp. strain B1 90 6.0–7.0 Marjaana et al. (1994)

Fusarium proliferatum 55 5.0–5.5 Badal (2002)

Pyrococcus furiosus 100 6.0 Bauer et al. (1999)

Pyrococcus furiosus 102 – Kengen et al. (1993)

Scytalidium thermophilum 65 6.0 Nazan and Zumrut (2000)

Streptomyces sp. strain S38 60 6.0 Georis et al. (2000)

Sulfolobus solfataricus 105 5.3 Nucci et al. (1993)

Teheromyces lanuginosus (wild and mutant) 60, 70 7.0, 6.7 Bakalova et al. (2002)

Teheromyces lanuginosus-SSBP 70–75 6.5 Johnson et al. (1999)

Thermoascus aurantiacus 50 5.0 Kalegoris et al. (1998)

Thermotoga maritima MSB8 92 6.2 Winterhalter and Liebl (1995)

Thermotoga maritima MSB8 95 6.0–7.5 Bronnenmeier et al. (1995)

Thermotoga maritima MSB8 75 6.2 Gabelsberger et al. (1993)Thermotoga neapolitana 95 6.0 Bok et al. (1998)

Thermotoga neapolitana 85 5.5 Bok et al. (1994)

Thermotoga neapolitana 102 5.5 Zverlov et al. (1996)

Thermotoga sp. strain FjSS3-B1 80 7.0 Ruthersmith and Daniel (1991)

Thermotoga sp. strain FjSS3-B1 105 6.8–7.8 Ruthersmith and Daniel (1991)

Thermotoga sp. strain FjSS3-B1 115 5.3 Simpson et al. (1991)

Thermotoga thermarum 80 6.6 Sunna et al. (1996a)




mode. The difference in the type of bond and the highly

ordered crystalline form of the compound between

starch and cellulose make cellulose more resistant to

digest and hydrolyze. The enzymes required for the

hydrolysis of cellulose include endoglucanases, exoglu-

canases and b-glucosidases (Matsui et al., 2000). While

cellulase is an endoglucanase that hydrolyses cellulose

randomly, producing oligosacaccharides, cellobiose and

glucose, exoglucanases hydrolyze b-1,4-DD-glucosidic

linkages in cellulose releasing cellobiose from the non-

reducing end. On the other hand, b-glycosidases of

thermophilic origin, which have received renewed at-

tention in the pharmaceutical industry hydrolyze cello-

biose to glucose.

In the current industrial processes, cellulolytic en-

zymes are employed in the color extractions of juices, in

detergents causing color brightening and softening, in

the biostoning of jeans, in the pretreatment of biomass

that contains cellulose to improve nutritional quality of

forage and in the pretreatment of industrial wastes(Buchert et al., 1997; Niehaus et al., 1999; Bhat, 2000;

Nakamura et al., 2001; Van wyk et al., 2001; Zhou et al.,

2001). In order to attack the native crystalline cellulose,

which is water insoluble and occurs as fibers of densely

packed structures, however, thermostable cellulases

active at high temperature and high pH are required.

Thermostable cellulases of archaeal origin include

those isolated from Pyrococcus furiousus (Kengen et al.,

1993) and Pyrococcus horikoshi (Ando et al., 2002).

While the latter has an optimum temperature of 97 °C,

the enzyme from Pyrocuccus furiousus has shown opti-

mal activity at 102–105 °C (Table 6). Sulfolobus solfa-

taricus MT4, Sulfolobus acidocaldarius and Sulfolobus

shibatae were also described as producers of b-gucosid-

ases (Grogan, 1991). From Thermotoga maritema MSB8

optimally active cellulase acting at 95 °C and between

pH 6.0 and 7.0 was reported (Bronnenmeier et al., 1995).

Other endocellulases (CelA and CelB) from Thermotoga

neapolitana, were purified and characterized (Bok et al.,

1998). Optimal pH for CelA is 6.0 at 95 °C, and the pH

range of CelB is broad (6.0–6.6) at 106 °C. CelA, with

the ability to hydrolyze microcrystalline cellulose, was

isolated from the extremely thermophilic bacterium

Anaerocellum thermophilum (Zverlov et al., 1998) and

maximal activity of this enzyme was observed at pH

5.0–6.0 and 85–95 °C. A highly thermostable cellobiose

(115 °C at pH 6.8–7.8) was also produced from Ther-

motoga sp. FjSS3-B1 (Ruthersmith and Daniel, 1991,

1992).

It is obvious that, the successful utilization of cellu-

lose is dependent on the development of economically

feasible technologies for the production of cellulase. The

biopolishing process of cotton in the textile industry, for

example, requires cellulase stable at high temperature

close to 100 °C (Ando et al., 2002). Presently used en-

zymes for this purpose, however, are active only at 50–

55 °C. Cellulase production is also found to be the most

expensive step during ethanol production from cellulosic

biomass, and accounted for approximately 40% of the

total cost (Spano et al., 1975). In the food industry,degradation of cellulose by acids is still unsatisfactory

and results in the decomposition of the sugars. Finally,

even though many cellulolytic enzymes of thermophillic

origin are known their function under physiological

condition remains unclear.

8. Thermoactive chitinases

Chitin, the second most abundant natural biopolymer

after cellulose, consists of a linear b-1,4-homopolymer

of N

-acetylglucoseamine residues. In nature, it is usuallyfound attached to other polysaccharides and proteins.

The covering layer of insects, cell walls of various fungi

and crab and shrimp wastes are the main sources of

chitin (Majeti and Kumar, 2000; Nwe and Stevens,

2002; Suntornsuk et al., 2002). The major applications

of chitosan include wastewater clearing, preparation of

cosmetics, paper production, textile finishes, photo-

graphic products, cements, heavy metal chelating agents

Table 6

Source microorganisms and properties of thermostable cellulasesOrganism Enzyme properties References


Anaerocellu thermophilum 85–90 5.0–6.6 Zverlov et al. (1998)

Bacillus subtilis 65–70 5.0–6.5 Mawadza et al. (2000)

Pyrococcus furiosus 102–105 – Kengen et al. (1993)

Pyrococcus horicoshi 97 – Ando et al. (2002)

Rhodothermus marinus 95 6.5–8.0 Hreggvidsson et al. (1996)

Thermotoga maritema MSB8 95 6.0–7.0 Bronnenmeier et al. (1995)

Thermotoga neapoltana

(EndocellulaseA)

95 6.0 Bok et al. (1998)

Thermotoga neapoltana

(EndocellulaseB)

106 6.0–6.6 Bok et al. (1998)




and for medical and veterinary purposes (Spindler

et al., 1990; Georgopapadakou and Tkacz, 1995;

Hirano, 1996; Cohen and Chet, 1998; Majeti and

Kumar, 2000).

The production of chitosan from chitin involves a

deacetylation reaction that is done using 40% sodium

hydroxide solution. This is a highly corrosive stream

which is difficult to control. Attempts are thus being

made to do this conversion using deacetylase enzyme.

Perhaps a thermostable deacetylase enzyme will help in

increasing yields and conversion rates and allow reuse of

the enzyme.

Endo-acting chitin hydrolase chitinaseA, the exo-

acting hydrolase chitinaseB and N -acetyl-DD-glucosea-

minidase are responsible for chitin degradation (Kenji

et al., 1994). However, chitin is not easily accessible to

chitinases and chitin deacetylases. Evidences from X-ray

diffraction studies have shown that chitin is a highly

ordered crystalline structure and does not dissolve in

water (Roberts, 1992). This property of chitin has shownthe need for thermostable enzymes.

The thermophilic organisms Bacillus licheniformis X-

7u (Takayanagi et al., 1991), Bacillus sp. BG-11 (Bharat

and Hoondal, 1998) and Strteptomyces thermoviol-

aceus OPC-520 were reported to be the major sources

of chitinases (Tsujibo et al., 1995). Chitinase and N -

acetyl-glucosaminidase were produced from the ex-

treme thermophilic anaerobic arachaeon Thermococcus

chitinophagus (Huber et al., 1995). A thermophilic bac-

terium strain producing three different endochitinases in

its culture fluid was isolated from chitin-containing

compost (Kenji et al., 1994). The strains belonged to the

genus Bacillus and three isoforms of endochitinases (L,

M and S) were purified and characterized. They had

different molecular masses (71, 62 and 53 Kda) and

showed temperature optima of 75, 65 and 75 °C at a pH

of 6.5, 5.5 and 5.5 respectively. The iso-electric points

were 5.3, 4.8, and 4.7. Thermostable exochitinases were

also isolated from Bacillus stearothermophilus CH-4,

isolated from a compost of organic solid wastes (Kenji

et al., 1998).

The basic parts of shrimp and prawns used for con-

sumption are the muscle tissues. The resulting waste of

the products, thus, is vast (Healey et al., 1994). In some

developing countries it was reported that more than 2.5million tones of shrimp and prawns are produced per

year (Anon., 1996). Therefore, there is a high need to

develop methods of producing value added products

from this waste. As the major way to exploit the po-

tential from this waste is the production of chitin and its

derivative chitosan, improved and economically feasible

chemical, enzymatic and novel biotechnological con-

version bioprocesses are required. Chitin together with

its derivatives can also be used as agrochemicals (Kenji

et al., 1998). However, there is only little effort to clarify

its utilization in the food industry.

9. Thermostable proteases

Proteases, which are generally classified into two

categories (exopeptidases, that cleave off amino acids

from the ends of the protein chain and endopeptidases,

which cleave peptide bonds within the protein) are be-

coming major industrial enzymes, and constitute more

than 65% of the world market (Rao et al., 1998). These

enzymes are extensively used in the food, pharmaceuti-

cal, leather and textile industries (Cowan, 1996; Fan

et al., 2001; Mozersky et al., 2002). The applications will

keep increasing in the future as will the need for stable

biocatalysts capable of withstanding harsh conditions of

operation.

Relative ease of isolation of Bacilli from diverse

sources has made these organisms the focus of attention

in biotechnology (Johnevelsy and Naik, 2001). So far,

however, few thermophilic Bacillus sp. that produce

proteases have been isolated, the earliest isolate being

Bacillus stearothermophilus (Salleh et al., 1977) which isstable at 60 °C. Another Bacillus sp. has produced a

thermostable protease that has an optimum activity at

60 °C (Razak et al., 1993), while a different Bacillus

stearothermophilus sp. produced an alkaline and ther-

mostable protease which is optimally active at 85 °C

(Razak et al., 1995, 1997; Rahman et al., 1994). A spe-

cies of Bacillus stearothermophilus TP26 that has been

isolated produces an extra cellular protease having an

optimum temperature of 75 °C (Gey and Unger, 1995).

Enhancement of protease activity excreted from Bacillus

stearothermophilus had also been possible using eco-

nomical chemical additives in the proteolysis reactions

involved in waste activated sludge (Kim et al., 2002a,b).

In a chemically defined medium, thermophilic and al-

kaliphilic Bacillus sp. JB-99 was also reported to pro-

duce thermostable alkaline proteases (Johnevelsy and

Naik, 2001). Dominant producers of proteases in fact,

are the microorganisms of the genera Pyrococcus,

Thermococcus and Staphylothermus (Table 7). Ex-

tremely thermostable serine proteases are produced by

the hyperthermophilic archaeum Desulfurococcus strain

(Hanazawa et al., 1996), and thermostable metallopro-

teases are reported from a gram-negative thermophilic

bacterium (Pawinee and Wipapat, 1996).

Major area of focus in the future concerning theproduction of proteases is the optimization of media

(Johnevelsy and Naik, 2001). Synthetic media have a

great advantage over complex media in that consistency

of processes and production is enhanced through

avoiding the variability of complex substrates. Thus,

synthetic media provides better control and monitoring,

improved product recovery and quality, and simplified

purification systems (Zhang and Greasham, 1999). It is

for this reason that reports on the production of pro-

tease enzymes using synthetic media are available re-

cently (Mao et al., 1992; Ferrero et al., 1996).




There is considerable current interest on the explo-

ration of proteases that can catalyze reactions in cold

water (Demirijan et al., 2001). This will allow their use in

detergents which can be used in normal tap water

without the requirement for increasing the temperature

of the water. The search for such enzymes is very much a

challenge at this time.

10. Heat stable lipases

Lipases of microbial origin are the most versatileenzymes and are known to bring about a range of bio-

conversion reactions (Vulfson, 1994), which includes

hydrolysis, interesterification, esterification, alcoholysis,

acidolysis and aminolysis (Jaeger et al., 1994; Pandey

et al., 1999; Nagao et al., 2001; Kim et al., 2002a,b).

Their unique characteristics include substrate specifi-

city, stereospecificity, regioselectivity and ability to cata-

lyze a heterogeneous reaction at the interface of water

soluble and water insoluble systems (Brockman and

Borgstorm, 1984; Jaeger and Reetz, 1998).

The esters produced play a relevant role in the food

industry as flavor and aroma constituents (Gandhi et al.,

1995; Pandey et al., 1999). Whereas long chain methyland ethyl esters of carboxylic acid moieties provide

valuable oleo chemical species that may function as fuel

for diesel engines, esters of long chain carboxylic acid

and alcohol moieties (waxes) have applications as lub-

ricants and additives in cosmetic formulations (Linko

et al., 1994). Other applications include the removal of

the pitch from pulp produced in the paper industry, for

the hydrolysis of milk fat in the dairy industry, removal

of non-cellulosic impurities from raw cotton before

further processing into dyed and finished products, drug

formulations in the pharmaceutical industry and in the

removal of subcutaneous fat in the leather industry

(Pandey et al., 1999; Traore and Buschle-Diller, 2000).

A biodiesel was derived from vegetable oils using im-

mobilized Candida antarctica lipase (Shimada et al.,

1999).

Most of the industrial processes in which lipases are

employed function at temperatures exceeding 45 °C. The

enzymes, thus, are required to exhibit an optimum

temperature of around 50 °C (Sharma et al., 2002).

According to some reports there are fats exhibiting

higher melting points and which are able to inhibit

enzymatic reactions at a low temperature (Dong-Woo

et al., 1999). Some enzymatic processes for the physical

refining of seed oils have four distinct requirements.

These include pH of 5.0 and optimal temperature of

around 65 °C, adding an enzyme solution and enzyme

reaction followed by the separation of the lysophos-

phatide from the oil at about 75 °C (Klaus, 1998). These

reactions, therefore, are enhanced through the utiliza-

tion of thermo-tolerant lipases.

Lipases are of widespread occurrence throughout the

earthÕs flora and fauna. More abundantly, however, they

are found in bacteria, fungi and yeasts (Wu et al., 1996).

Several Bacillus sp. were reported to be the main source

of lypolytic enzymes (Kim et al., 1994; Schmidt et al.,1994; Luisa et al., 1997). While most of these enzymes

are active at a temperature of 60 °C and pH of 7.0,

lipases from Bacillus thermoleovorans and a thermo-

philic Rhizopus oryzae strain can moderately function at

extreme pH and temperature values (Dong-Woo et al.,

1999; Abel et al., 2000). The pH and temperature optima

for the catalytic activity of some thermostable lipases

are given in Table 8.

Reports of thermostable lipases from archaeal ori-

gin, however, are very few. Phospholipase A2 which

was secreted from the archaea Pyrococcus horikoshii

Table 7

Source microorganisms and properties of thermostable proteolytic enzymes



Bacillus brevis 60 10.5 Banerjee et al. (1999)

Bacillus licheniformis 70 9.0 Manchini and Foretina (1998)

Bacillus stearothermophilus 60 – Salleh et al. (1977)

Bacillus stearothermophilus 85 – Rahman et al. (1994)Bacillus sp. JB-99 80 6–12 Johnevelsy and Naik (2001)

Bacillus stearothermophilus TP26 75 – Gey and Unger (1995)

Bacillus sp. no. AH-101 80 12.0–13.0 Takami et al. (1989)

Bacillus thermoruber 45 9 Manchini et al. (1988)

Pyrococcus sp. KODI 100 7 Fujiwara et al. (1996)

Staphylothermus marinus – 9 Mayer et al. (1996)

Thermoacidophiles

(archeal and bacterial origin)

60–70 7.0–8.5 Kocabiyik and Erdem (2002)

Thermococcus aggreganes 90 7.0 Klingberg et al. (1991)

Thermococcus celer 95 7.5 Klingberg et al. (1991)

Thermococcus litoralis 85 8.5 Klingberg et al. (1991)

Thermotoga maritema 95 9.5 Klingberg et al. (1991)




(Yan et al., 2000) was involved in crude oil refining.

This enzyme was found to optimally react at 95 °C andpH of 7.0. Phospholipase A2 has a better performance in

the degumming processes of oil refineries and thereby re-

duces wastewater problems and running costs (Klaus,

1998).

Among the desirable characteristics that commer-

cially important lipases should exhibit, alkali tolerance

and thermostability are the main (Kulkurani and Gadre,

1999). To meet this end, there is a continuous search for

sources of highly active lipolytic enzymes with specific

stability to pH, temperature, ionic strength and organic

solvents (Lee and Rhee, 1993). Although, few lipases

exist which are able to operate at 100 °C, their half-lives

are reported to be short (Rathi et al., 2000).

The other problem associated with the production of

lipases and which requires improvements is its secretion

at the late stage of growth (Tan and Gill, 1985). Prote-

ases secreted in the mean-time either change the prop-

erties of the lipase produced or degrade it (Cordenons

et al., 1996). Perhaps it is for this reason that only few

lipases can be obtained in industrial quantities (Wu

et al., 1996). Hence the need for a more stable lipase

enzyme produced in large quantities.

11. Thermostable DNA polymerases

The polymerase chain reaction (PCR) process has led

to a huge advance in genetic engineering due to its ca-

pacity to amplify DNA. The three successive steps in

this process include denaturation or melting of the DNA

strand (separation) obtained at a temperature of 90–95

°C, renaturation or primer annealing at 55 °C followed

by synthesis or primer extension at around 75 °C (Mullis

et al., 1986; Erlich et al., 1988; Saiki et al., 1988). De-

velopment in this process has been to a large extent

facilitated by the availability of thermostable DNA

polymerases, which catalyse the elongation of the primer

DNA strand (Mullis and Faloona, 1987).In earlier PCR procedures, DNA polymerases which

were isolated from Escheria coli were utilized (Mullis

et al., 1986; Erlich et al., 1988). These enzymes, how-

ever, lost their enzymatic activities at elevated temper-

atures and, thus, adding a new polymerase enzyme

after each cycle following the denaturation and primer

hybridization steps was necessary. This process made

the thermal cycling a time-consuming and costly pro-

cedure.

Taq polymerase from the bacterium Thermus aquat-

icus was the first thermostable DNA polymerase char-

acterized (Chien et al., 1976; Kaledin et al., 1980).

Repeated exposure to 98 °C in a reaction buffer had little

effect on the enzyme activity and significant activity re-

mained after exposure to 99 °C. Although, Taq poly-

merase exhibits a 50 –30 exonuclease activity, a 30 –50

exonuclease activity was not detected. Thus, the base

insertion fidelity is low as the enzyme is unable to correct

misincorporated nucleotides (Longley et al., 1990; Ling

et al., 1991). It had also been possible to determine the

error rates of some polymerases in terms of base pairs

(Alkami Biosystems, 1999).

Table 9 shows a variety of thermostable polymerases

with 30 –50-exonuclease-dependent proof reading activity

required from a high fidelity polymerase. Optimizationof the PCR procedure can be done by mixing a standard

polymerase, such as Taq polymerase with high fidelity

polymerases (Pfu, Vent and Deep Vent) (Ling et al.,

1991).

While the PCR process is developing rapidly through

the invention of better instruments (Mariella, 2001), lack

of fidelity has remained to be a serious challenge. The

five distinct activities in which errors can occur are the

rate of phosphodiester bond formation, the binding of

dNTP by the polymerase, the rate of pyrophospahte

release, contamination after misincorporation and the

Table 8

Source microorganisms and properties of thermostable lipases



Bacillus acidocaldariusa (esterase) 70 – Manco et al. (1998)

Bacillus sp. RSJ-1 50 8.0–9.0 Sharma et al. (2002)

Bacillus strin J 33a 60 8.0 Nawani et al. (1998)

Bacillus stearothermophilusa 68 – Gupta et al. (1999)Bacillus thermocatenletusa 60–70 8.0–9.0 Klibanov (1983)

Bacillus thermoleovorans ID-1 70–75 7.5 Dong-Woo et al. (1999)

Geobacillus sp. 70 9.0 Abdel-Fattah (2002)

Pseudomonas sp. 65 9.6 Kulkurani and Gadre (1999)

Pseudomonas sp. 90 11.0 Rathi et al. (2000)

Pyrobaculum calidifontis 90 – Hotta et al. (2002)

Pyrococcus furiosusa (esterase) 100 – Ikeda and Clark (1998)

Pyrococcus horikoshii 97 5.6 Ando et al. (2002)

Pyrococcus horikoshii 95 7.0 Yan et al. (2000)

a Active at water immiscible solvents.




30 –50-exonuclease proof reading capacity of the enzyme

(Alkami Biosystems, 1999). Some issues that nowadays

have stimulated scientists in the area thus, are im-

provements required in the fidelity of the PCR, devel-opment of strategies for more economical use of the

enzymes and ensuring rapid multiplication from fewer

strands. A thermostable DNA polymerase having these

characteristics will indeed improve the results obtained

by PCR machine.

12. Molecular cloning of thermophilic genes into meso-

philic hosts

Genetic and protein engineering are the modern

techniques for the commercial production of enzymes of improved stability to high temperatures, extremes of

pH, oxidizing agents and organic solvents. Cloning and

expression of genomic information available in a ther-

mophile in a suitable and faster growing mesophilic host

has also provided possibilities of producing the speci-

fic thermostable enzyme required for a particular bio-

transformation process (Blackebrough and Birch, 1981;

William and Dennis, 1988; James, 1995; Ikeda and

Clark, 1998; Hough and Danson, 1999).

Genes encoding a-amylase (amyA), pullulanase

( pulA), maltodextrin phosphrylase (agpA) and a-glu-

cosidase (aglA) were identified in the gene library of

Thermotoga maritema (Bibel et al., 1998). The enzymesproduced after expression in Escheria coli are reported

to demonstrate extreme thermostability. Pyrococus fu-

riosus a-amylase gene, which was cloned by activity

screening in Escheria coli was reported to be optimally

active at 100 °C, pH 5.5–6.0 and not to require Ca2þ for

its activity (Dong et al., 1997a). Genes encoding pullu-

lanases from Pyrococus furiosus (Dong et al., 1997b) and

Ferividobacterium pennavorans (Bertoldo et al., 1999),

Thermotoga neapolitana genes encoding for xylanases

(Velikodvorskaya et al., 1997), Protease genes from

Pyrobaculum aerophilum and Bacillus stearothermophilus

(Volkl et al., 1995; Kubo and Imanaka, 1988), lipases

from Bacillus thermocatenulatus (Schmidt et al., 1994),

esterases from Bacillus licheniformis (Alvarez-macarie

et al., 1999) and cellulase genes from the archaon Py-rococus furiosus (Bauer et al., 1999) were expressed from

an Escheria coli recombinant strain. The production of

cellulase enzyme was further enhanced through DNA

recombinant technology by encoding the genes for cel-

lulase in various species of Clostridium (Beguin, 1992).

In addition to this, the nucleotide sequence and the

cloning of CelA and CelB genes from C. thermocellum

were studied and reported (Bergquist et al., 1992).

Cloning and expression of chitinase genes was also

successfully done (Duffner et al., 1996; Chernin et al.,

1997). In cases where a relevant gene of a thermostable

enzyme has been identified and once the gene has been

transferred into a mesophilic host the resultant enzymeproduction can be facilitated through appropriate in-

cubation processes.

13. Conclusion

Recent investigations have demonstrated that ex-

tremophilic archaea, bacteria and fungi have colonized

environments that were believed to be inhospitable for

survival. Their true diversity in fact, is not yet been fully

explored. The thermostable enzymes isolated from these

organisms have just started providing conversions underconditions that are appropriate for industrial applica-

tions. The conditions required by these thermostable

enzymes which bring about specific reactions not pos-

sible by chemical catalysts are still mild and environ-

mentally benign, as compared to the temperatures and

pressures required for chemical conversions. Thus, with

the availability of thermostable enzymes a number of

new applications in the future are likely. Although, be-

lieved to provide tremendous economical benefits, pro-

duction of the enzymes to the level required by the

industries has remained a challenge.

Table 9

Thermostable DNA polymerases in PCR

Polymerases Organism PCR reactions References

BstI pol Bacillus stearothermophilus HF Mead et al. (1991)

Deep Vent Pol Pyrococcus sp. GB-D LF Cline et al. (1996)

Pfu pol Pyrococcus furiosus LF Lundberg et al. (1991)

Pwo pol Pyrococcus woesei LF Frey and Suppman (1995)

Taq pol I Thermus aquaticus HF Jones and Foulkes (1989)Tfi pol Thermus filiformis NI Perler et al. (1996)

Tfl pol Thermus flavus NI Kaledin et al. (1980)

Tth pol Thermus thermophilus HF Myers and Gelfand (1991),

Pantazaki et al. (2002)

Tma pol Thermotoga maritema LF Bost et al. (1994)

Vent pol Thermococcus litoralis LF Perler et al. (1996)

HF and LF, high and low fidelity; NI, not identified.




References

Abdel-Fattah, Y., 2002. Optimization of thermostable lipase produc-

tion from a thermophilic Geobacillus sp. using Box–Behnken

experimental design. Biotechnol. Lett. 24, 1217–1222.

Abel, H., Marie, D., Nathalie, R., Danielle, D., Louis, S., Louis, C.,

2000. Purification and characterization of an extracellular lipase

from a thermophilic Rhizopus oryzae strain isolated from palm

fruit. Enzyme Microb. Technol. 26, 421–430.

Aguilar, G., Morlon-Guyot, J., Trejo-Aguilar, B., Guyot, J.P., 2000.

Purification and characterization of an extracellular alpha-amylase

produced by Lactobacillus manihotivorans an amylolytic lactic acid

bacterium. Enzyme Microb. Technol. 27, 406–413.

Alkami Biosystems, 1999. Alkami quick guide TM for PCR. Alkami

Biosystems Inc., USA.

Alvarez-macarie, E., Augier-Magro, V., Baratti, J., 1999. Character-

ization of a thermostable esterase activity from the moderate

thermophile Bacillus licheniformis. Biosci. Biotechnol. Biochem. 63,

1865–1870.

Andersson, L., Rydberg, U., Larsson, H., Andersson, R., Aman, P.,

2002. Preparation and characterisation of linear dextrins and their

use as substrates in in vitro studies of starch branching enzymes.

Carbohyd. Polym. 47, 53–58.

Ando, S., Ishida, H., Kosugi, Y., Ishikawa, K., 2002. Hyperthermo-

stable endoglucanase from Pyrococcus horikoshi . Appl. Environ.

Microbiol. 68, 430–433.

Anon., 1996. World Review of Fisheries and Aquaculture. FAO,

United Nations.

Antranikian, G., Herzberg, C., Gottschalk, G., 1987. Production of

thermostable a-amylase, pullulanase and a-glucosidase in contin-

uous culture by a new Clostridium isolate. Appl. Environ. Micro-

biol. 53, 1668–1673.

Ashita, D., Gupta, J., Khanna, S., 2000. Enhanced production,

purification and characterization of novel cellulase-poor thermo-

stable, alkali tolerant xylanase from Bacillus circulans AB 16. Proc.

Biochem. 35, 849–856.

Badal, C., Gwo-Jenn, S., Kailash, C., Lloyd, W., Gregory, Z., 1989.

New thermostable a-amylase-like pullulanase from thermophilc

Bacillus sp. 3183. Enzyme Microb. Technol. 11, 760–764.

Badal, C., 2002. Production, purification and properties of xylanase

from a newly isolated Fusarium proliferatum. Proc. Biochem. 37,

1279–1284.

Bahrami, A., Shojaosadati, S., Mahbeli, G., 2001. Biodegradation of

dibenzothiophene by thermophilic bacteria. Biotechnol. Lett. 23,

899–901.

Bakalova, N., Petrova, S., Atev, A., Bhat, M., Kolev, D., 2002.

Biochemical and catalytic properties of endo-1,4-b-xylanases from

Thermomyces lanuginosus (wild and mutant strains). Biotechnol.

Lett. 24, 1167–1172.

Banerjee, V., Saani, K., Azmi, W., Soni, R., 1999. Thermostable

alkaline protease from Bacillus brevis and its characterization as a

laundry additive. Proc. Biochem. 35, 213–219.

Bataillon, M., Cardinali, A.-P., Castillon, N., Duchiron, F., 2000.Purification and characterization of a moderately thermostable

xylanase from Bacillus sp. strain SPS-0. Enzyme Microb. Technol.

26, 187–192.

Bauer, M., Driskil, L., Callen, W., Snead, M., Mathur, E., Kelly, R.,

1999. An endoglucanase EglA, from the hyperthermophilic archa-

eon Pyrococcus furiosus hydrolyzes a-1,4 bonds in mixed linkage

(1!3), (1!4)-b-DD-glucans and cellulose. J. Bacteriol. 181, 284–290.

Becker, P., 1997. Determination of the kinetic parameters during

continuous cultivation of the lipase producing thermophile Bacillus

sp. IHI-91 on olive oil. Appl. Microb. Biotechnol. 48, 184–190.

Beguin, P., 1992. Molecular biology of cellulose degradation. In:

Herbert, R., Sharp, R. (Eds.), Molecular Biology and Biotechnol-

ogy of Extremophiles. Chapman and Hall, NY.

Bergquist, P., Love, D., Croft, J., Streiff, M., Daniel, R., Morgan, H.,

1992. Cloning of genes involved in cellulose hydrolysis. In: Herbert,

R., Sharp, R. (Eds.), Molecular Biology and Biotechnology of

Extremophiles. Chapman and Hall, NY.

Bertoldo, C., Duffner, F., Jorgensen, P., Antrakianin, G., 1999.

Pullulanase type I from Ferividobacterium pennavorans Ven5:

cloning, sequencing, expression of the gene and biochemical

characterization of the recombinant enzyme. Appl. Environ.

Microbiol. 65, 2084–2091.Bharat, B., Hoondal, G., 1998. Isolation, purification and properties of

thermostable chitinase from an alkalophilic Bacillus sp. BG-11.

Biotechnol. Lett. 20, 157–159.

Bhat, M., 2000. Cellulases and related enzymes in biotechnology.

Biotechnol. Adv. 18, 355–383.

Bibel, M., Bretel, C., Gosslar, U., Kriegshauser, G., Liebl, W., 1998.

Isolation and analysis of genes for amylolytic enzymes of the

hyperthermophilic bacterium Thermotoga maritema. FEMS Mi-

crobiol. Lett. 158, 9–15.

Blackebrough, N., Birch, G., 1981. Enzymes of Food Processing.

Applied Science Publishers, London.

Bok, J., Goers, S., Eveleigh, D., 1994. Cellulase and xylanase systems

of Thermotoga neapolitana. ACS Symp. Ser. 566, 54–65.

Bok, J., Dienesh, A., Yernool, D., Eveleigh, D., 1998. Purification,

characterization and molecular analysis of thermostable cellulasesCelA and CelB from Thermotoga neapolitana. Appl. Environ.

Microbiol. 64, 4774–4781.

Bost, D., Stoffel, S., Landre, P., Lawyer, F., Akers, J., Abramson, R.,

Gelfand, D., 1994. Enzymatic characterization of Thermotoga

maritema DNA polymerase and a truncated form, Ultma DNA

polymerase. FASEB J., A1395.

Breccia, J., Sineriz, F., Baigori, M.D., Guillermo, R.C., Hatti-Kaul,

R., 1997. Purification and characterisation of a thermostable

xylanase from Bascillus amyloliquefaciens. Enzyme Microb. Tech-

nol. 22, 42–49.

Brockman, H., Borgstorm, B., 1984. Lipases. Elsevier, Amsterdam.

Bronnenmeier, K., Kern, A., Libel, W., Staudenbauer, W., 1995.

Purification of Thermotoga maritema enzymes for the degradation

of cellulose materials. Appl. Environ. Microbiol. 61, 1399–1407.

Brown, S., Kelly, R., 1993. Characterization of amylolytic enzymeshaving both a 1,4 and a 1,6 hydrolytic activity from the

thermophilic archaea Pyrococcus furiosus and Thermococcus lito-

ralis. Appl. Environ. Microbiol. 59, 2614–2621.

Brown, C., 1987. Enzyme technology. In: Wilkinson, J. (Ed.),

Introduction to Biotechnology. Black Well Scientific, Oxford.

Buchert, J., Pere, J., Oijusluoma, L., Rahkamo, L., Viikari, L., 1997.

Cellulases––tools for modification of cellulosic materials. In:

Niches in the World of Textiles World Conference of the Textile

Institute, Manchester, England, pp. 284–290.

Burrows, W., 1973. Text Book of Microbiology. W.B. Saunders

Company, Ontario.

Canganella, F., Andrade, C., Antranikian, G., 1994. Characterization

of amylolytic and pullulytic enzymes from thermophilic archaea

and from a new Ferividobacterium species. Appl. Microb. Biotech-

nol. 42, 239–245.

Carlsen, M., Nielsen, J., Jorgensen, S.B., Zangirolami, T.C., 2002.

Growth and enzyme production during continuous cultures of a

high amylase-producing variant of Aspergillus oryzae. Braz. J.

Chem. Eng. 19, 55–67.

Chen, C., Adolphson, R., Dean, F., Eriksson, K., Adamas, M.,

Westpheling, J., 1997. Release of lignin from kraft pulp by a

hyperthermophilic xylanase from Thermotoga maritema. Enzyme

Microb. Technol. 20, 39–45.

Chernin, L.S., de la Fuente, L., Sobolev, V., Haran, S., Vorgias, C.E.,

Oppenheim, B.A., Chet, I., 1997. Molecular cloning, structural

analysis and expression in E. coli of a chitinase gene from

Enterobacter agglomerans. Appl. Environ. Microbiol. 63, 834–839.




Chien, A., Edgar, D., Trela, J., 1976. Deoxyribonucleic acid polymer-

ase from the extreme thermophilile Thermus aquaticus. J. Bacteriol.

127, 1550–1557.

Chitradon, L., Mahakhan, P., Bucke, C., 2000. Oligosaccharide

synthesis by reversed catalysis using alpha-amylase from Bacillus

licheniformis. J. Mol. Catal. B: Enzym. 10, 273–280.

Clarke, J., Rixon, J., Ciruela, A., Gilbert, H., Hazlewood, G., 1997.

Family-10 and Family-11 xylanases differ in their capacity to

enhance the bleachability of hardwood and softwood paper pulps.Appl. Microb. Biotechnol. 48, 177–183.

Cline, J., Braman, J., Hogrefe, H., 1996. PCR fidelity of Pfu DNA

polymerase and other DNA polymerases. Nucleic Acids Res. 24,

3546–3551.

Cohen, K.R., Chet, I., 1998. The molecular biology of chitin digestion.

Curr. Opin. Biotechnol. 9, 270–277.

Cordenons, A., Gonzalez, R., Kok, R., Hellingwerf, K., Nudel, C.,

1996. Effect of nitrogen sources on the regulation of extracellular

lipase production in Acinetobacter calcoaceticus strains. Biotech-

nol. Lett. 18, 633–638.

Cowan, D., 1996. Industrial enzyme technology. Trends Biotechnol. 14

(6), 177–178.

Crab, W., Mitchinson, C., 1997. Enzymes involved in the processing of

starch to sugars. Trends Biotechnol. 15, 349–352.

Daniel, L., 1979. Fermentation and Enzyme Technology. John Willeyand Sons, NY.

Danson, M., Hough, D., Lunt, G., 1992. The Archaebacteria:

Biochemical and Biotechnology. Portland, London.

De Mot, R., Andries, K., Verachtert, H., 1984a. Comparative study of

starch degradation and amylase production by Ascomycetes yeast

species. Syst. Appl. Microbiol. 5, 106–118.

De Mot, R., Van Oudendijck, E., Verachert, H., 1984b. Production of

extracellualar debranching activity by amylolytic yeasts. Biotech-

nol. Lett. 6, 581–586.

De Rosa, M., Morana, A., Riccio, A., Gambacorta, A., Trincone, A.,

Incani, O., 1994. Lipids of the archaea: a new tool for bioelec-

tronics. Biosens. Bioelectr. 9, 669–675.

Dekker, R., Linder, W., 1979. Bioconversion of hemicellulose: aspects

of hemicellulase production by Trichoderma reesi QM9414 and

enzymic sacchrification of hemicellulose. S. Afr. J. Sci. 75, 65–71.Demirijan, D., Moris-Varas, F., Cassidy, C., 2001. Enzymes from

extremophiles. Curr. Opin. Chem. Boil. 5, 144–151.

Dhillon, A., Khanna, S., 2000. Production of a thermostable alkali-

tolerant xylanase from Bacillus circulans AB16 grown on wheat

straw. World J. Microbiol. Biotechnol. 27, 325–327.

Diane, W., Stephan, R., Wolfgang, Z., 1997. Application of thermo-

stable enzymes for carbohydrate modification. In: Contribution of

the Fourth International Workshop on Carbohydrate as Organic

Raw Materials, March 20–21, 1997. WUV-Universitatverlag,

Vienna.

Dijkhuizen, L., Van der Maarel, M., Van der Veen, B.U., 2002.

Properties and applications of starch-converting enzymes of the

alpha-amylase family. J. Biotechnol. 94, 137–155.

Dominguez, R., Souchon, H., Spinelli, S., Dauter, Z., Wilson, K.S.,

Chauvaux, S., Beguin, P., Alzari, P.M., 1995. A common protein

fold and similar active site in two distinct families of beta-

glycanases. Nat. Struct. Biol. 2, 569–576.

Dong, G., Vieille, C., Savachenko, A., Zeikus, G., 1997a. Cloning,

sequencing, and expression of the gene encoding extracellular a-

amylase from Pyrococcus furiosus and biochemical characterization

of the recombinant enzyme. Appl. Environ. Microbiol. 63, 3569–

3576.

Dong, G., Vieille, C., Savachenko, A., Zeikus, G., 1997b. Cloning,

sequencing, and expression of the gene encoding amylopullulanase

from Pyrococcus furiosus and biochemical characterization of the

recombinant enzyme. Appl. Environ. Microbiol. 63, 3577–3584.

Dong-Woo, L., You-Seok, K., Ki Jun, K., Byung-Chan, K., Hak-

Jong, C., Doo-sik, K., Maggy, T., Yu-ryang, P., 1999. Isolation

and characterisation of thermophilic lipase from Bacillus thermo-

leovorans ID-1. FEMS Microbiol. Lett. 179, 393–400.

Duffner, F., Christodoulou, E., Vorgias, C.E., 1996. Expression and

correct folding of a thermostable chitinase in E. coli . In: First

International Congress on Extremophiles, 2–6 June. Estoril,

Portugal.

Eichler, J., 2001. Biotechnological uses of archaeal extremozymes.

Biotechnol. Adv. 19, 61–278.

Emmanuel, L., Stefan, J., Bernard, H., Abdel, B., 2000. Thermophilicarchaeal amylolytic enzymes. Enzyme Microb. Technol. 26, 3–14.

Eriksson, L., Heitmann, J., 1998. Applications of enzyme technology

in the paper industry. In: Proceedings of the Technical Association

of the Pulp and Paper Industry, vol. 3, Montreal.

Erlich, H., Gelfand, D., Saiki, R., 1988. Specific DNA amplification

product review. Nature 33, 461–462.

Erra-Pujada, M., Chang, F., Debeire, P., Duchiron, F., OÕDonohue,

M., 2001. Purification and properties of the catalytic domain of the

thermostable pullulanase type II from Thermococcus hydrother-

malis. Biotechnol. Lett. 23, 1273–1277.

Estelle, L., Ladrat, C., Ann, G., Georges, B., Francis, D., 1997.

Thermostable amylolytic enzymes of thermophilic microorganisms

from deep-sea hydrothermal vents. Animal Biology. CR Acad. Sci.

Paris 320, 893–898.

Everly, C., Alberto, J., 2000. Stressors, stress and survival: overview.Front. Bioscien. 5, 780–786.

Fan, Z., Zhu, Q., Dai, J., 2001. Enzymatic treatment of wool. J. Dong

Hua University (English edition) 18, 112–115.

Ferrero, M., Castro, G., Abate, M., Baigori, M., Sinerz, F., 1996.

Thermostable alkaline protease of Bacilus licheniformis MIR 29:

isolation, production and characterization. Appl. Microbiol.

Technol. 45, 327–332.

Fitter, J., Herrmann, R., Hauss, T., Lechner, R.E., Dencher, N.A.,

2001. Dynamical properties of alpha-amylase in the folded and

unfolded state: the role of thermal equilibrium fluctuations for

conformational entropy and protein stabilization. Physica B:

Condens. Matter. 301, 1–7.

Fogarty, W., Kelly, C., 1995. Amylases, amyloglucosidases and related

glucanases. In: Enzyme and Microbiol Technology. In: Enzyme

and Microbial Systems Involved in Starch Processing, vol. 17, pp.770–778.

Fogarty, M., 1983. Microbial Enzymes and Biotechnology. Applied

Science Publishers, England.

Fontes, C., Hazlewood, G.P., Morag, E., Hall, J., Hirst, B.H., Gilbert,

H.J., 1995. Evidence for a general role for non-catalytic domains in

xylanases from thermophilic bacteria. Biochem. J. 307, 151–158.

Franks, F., 1993. Protein Biotechnology: Isolation, Characterization,

and Stabilization. Humana, Totowa, NJ.

Fredrich, A., Antrakian, G., 1996. Keratin degradation by Fervido-

bacterium pennavorans, a novel thermophilic anaerobic species of

the order Thermotogales. Appl. Environ. Microbiol. 62, 2875–

2882.

Frey, B., Suppman, B., 1995. Demonstration of the expand PCR

systemÕs greater fidelty and higher yields with a lacI-based fidelty

assay. Biochemica 2, 34–35.

Fujiwara, S., Okuyama, S., Imanaka, T., 1996. The world of archea:

genome analysis, evolution and thermostable enzymes. Gene 179,

165–170.

Gabelsberger, J., Liebl, W., Scheleifer, K., 1993. Cloning and

characterization of b-galactoside and b-glucoside hydrolysing

enzymes of Thermotoga maritema. FEMS Microbiol. Lett. 109,

130–138.

Gandhi, N., Sawant, S., Joshi, J., 1995. Studies on the lipozyme-

catalyzed synthesis of butyl laurate. Biotechnol. Bioeng. 46, 1–12.

Gantlet, H., Duchiron, F., 1998. Purification and properties of a

thermoactive and thermostable pullulanase from Thermococcus

hydrothermalis , a hyperthermophilic archaeon isolated from a deep

sea hydrothermal vent. Appl. Microbiol. Biotechnol. 49, 770–777.




Georgopapadakou, N., Tkacz, J., 1995. The fungal cell wall as a drug

target. Trends Microbiol. 3, 98–104.

Georis, J., Giannotta, F., De Buyl, E., Granier, B., Frere, J., 2000.

Purification and properties of three endo-beta-1,4-xylanases pro-

duced by Streptomyces sp. strain S38 which differ in their capacity

to enhance the bleaching of kraft pulp. Enzyme Microb. Technol.

26, 177–183.

Gessesse, A., 1998. Purification and properties of two thermostable

alkaline xylanases from an alkaliphilic Bacillus sp. Appl. Environ.Microbiol. 64, 3533–3535.

Gey, M., Unger, K., 1995. Calculation of the molecular masses of two

newly synthesized thermostable enzymes isolated from thermoph-

illic microorganismas. J. Chromatogr. B 166, 188–193.

Groboillot, A., 1994. Immobilization of cells for application in the

food industry. Crit. Rev. Biotechnol. 14, 75–107.

Grogan, W., 1991. Evidence that b-galactosidase of Sulfolobus

solfactoricus is only one of several activities of a thermostable b-

DD-glycosidase. Appl. Environ. Microbiol. 57, 1644–1649.

Gupta, R., Gupta, K., Sexena, R., Khan, S., 1999. Bleach-stable,

alkaline protease from Bacillus sp. Biotechnol. Lett. 21, 135–138.

Hamilton, M., Kelly, C., Fogarty, W., 1999. Production and properties

of the raw starch-digesting a-amylase of Bacillus sp. IMD 435.

Proc. Biochem. 35, 27–31.

Hanazawa, S., Hoaki, T., Jannasch, H., Maruyama, T., 1996. Anextremely thermostable serine protease from hyperthermophilic

archaeum Desulfurococcus strain sy, isolated from a deep sea

hydrothermal vent. J. Mar. Biotechnol. 4, 121–126.

Hans, S., 1995. Enzymes in Food Processing, A Reprint from

Biotechnology, second, completely revised ed. Weinheim.

Haseltine, C., Rolfsmeier, M., Blum, P., 1996. The glucose effect and

regulation of a-amylase synthesis in the hyperthermophilic archae-

aon Sulfolobus solfataricus. J. Bacteriol. 178, 945–950.

Hashida, M., Bisgaard-Frantzen, H., 2000. Protien engineering of new

industrial amylases. Trends Glycosci. Glycotechnol. 12, 389–401.

Healey, M.G., Romo, C.R., Bustos, R., 1994. Bioconversion of marine

crustacean shell waste. Resour. Conserv. Recycl. 11, 139–147.

Henrissat, B., Bairoch, A., 1993. New families in the classification of

glycosyl hydrolases based on amino acid sequence similarities.

Biochem. J. 293, 781–788.Herbert, R., Sharp, R., 1992. Molecular Biology and Biotechnology of

Extremophiles. Chapman and Hall, NY.

Hirano, S., 1996. Chitin biotechnological applications. In: Raafat El-

Gewely, M. (Ed.), Biotechnogical Annual Review, vol. 2. Elsevier

Press.

Holst, O., Manelius, A., Krahe, M., Markl, H., Raven, N., Sharp, M.,

1997. Thermophiles and fermentation technology. Comp. Biochem.

Physiol. 118A, 415–422.

Hotta, Y., Ezaiki, S., Atomi, H., Imanaka, T., 2002. Extremely stable

and versatile carboxyl esterase from a hyperthermophilic archaeon.

Appl. Environ. Microbiol. 68, 3925–3931.

Hough, D., Danson, M., 1999. Extremozymes. Curr. Opin. Chem.

Boil. 3, 39–46.

Hreggvidsson, G.O., Kaiste, E., Holst, O., Eggertsson, G., Palsdottir,

A., Kristjansson, J.K., 1996. An extremely thermostable cellulase

from the thermophilic eubacterium Rhodothermus marinus. Appl.

Environ. Microbiol. 62, 3047–3049.

Huber, H., Stetter, K., 1998. Hyperthermophiles and their possible

potential in biotechnology. J. Biotechnol. 64, 39–52.

Huber, R., Stoohr, J., Hohenhaus, S., Rachel, R., Burgraff, S.,

Jannsch, H., Stetter, K., 1995. Thermococcus chitonophagus sp.

Nov., a novel chitin degrading, hyperthermophilic archeum from

the deep sea hydrothermal vent environment. Arch. Microbiol. 164,

255–264.

Ikeda, M., Clark, D., 1998. Molecular cloning of extremely thermo-

stable esterase gene from hyperthermophilic archaeon Pyrococcus

furiosus in Escherhia coli . Biotechnol. Bioeng. 57, 624–629.

Jacques, G., Frederic, D.L., Joste, L.B., Viviane, B., Bart, D.,

Fabrizio, G., Benoit, G., Jean-marie, F., 2000. An additional

aromatic interaction improves the thermostability and thermophi-

licity of a mesophilic family 11 xylanase: structural basis and

molecular study. Protein Sci. 9, 466–475.

Jaeger, K., Reetz, T., 1998. Microbial lipases from versatile tools for

biotechnology. TIBTECH. 16, 396–403.

Jaeger, K., Ransac, S., Dijktra, C., Colson, C., Heuvel, M., Misset, O.,

1994. Bacterial lipases. FEMS Microbiol. Rev. 15, 29–63.Jakob, K., 1989. Thermophilic organisms as sources of thermostable

enzymes. TIBTECH. 7, 349–353.

James, A., 1995. Is thermophily a transferable property in bacteria.

Crit. Rev. Microbiol. 21, 165–174.

Jean-Jacques, A., Gladys, L., Sadok, K., Jacques, C., 1987. Isolation

and characterization of thermophilic bacterial strains with inulin-

ase activity. Appl. Environ. Microbiol. 53, 942–945.

Jeayoung, K., Takashi, N., Ryu, S., 1989. Thermostable, raw-starch-

digesting amylase from Bacillus stearothermophilus. Appl. Environ.

Microbiol. 55, 1638–1639.

Jensen, B., Norman, B., 1984. Bacillus acidopullulyticus pullulanase

application and regulatory aspects for use in the food industry.

Proc. Biochem. 19, 129–134.

Jensen, B., Olsen, J., 1991. Physicochemical properties of a purified

alpha-amylase from the thermphilic fungus Thermomyces lanugi-nosus. Enzyme Microb. Technol. 14, 112–116.

Jin, F., Li, Y., Zhang, C., Yu, H., 2001. Thermostable a-amylase and

a-galactosidase production from the thermophilic and aerobic

Bacillus sp. JF strain. Proc. Biochem. 36, 559–564.

Johnevelsy, B., Naik, G., 2001. Studies on production of thermostable

alkaline protease from thermophilic and alkaliphilic Bacillus sp.

JB-99 in a chemically defined medium. Proc. Biochem. 37, 139–144.

Johnson, L., Larr, M., Suren, S., Balakrishna, P., 1999. Characteristics

of b-DD-xylanase from a thermophilic fungus, Thermomyces lanu-

ginosus-SSBP. Biotechnol. Appl. Biochem. 30, 73–79.

Jones, M., Foulkes, N., 1989. Reverse transcription of mRNA by

Thermus aquaticus DNA polymerase. Nucleic Acids Res. 17, 8387–

8388.

Kaledin, A., Sliusarenko, A., Gorodetski, S., 1980. Isolation and

properties of DNA polymerase from extreme thermophilic bacteriaThermus aquqticus YT-1. Biochimiya 45, 644–651.

Kalegoris, E., Christakopoulos, D., Kekos, D., Macris, B., 1998.

Studies on the solid-state production of thermostable endoxylan-

ases from Thermoascus aurantiacus: characterization of two

isoenzymes. J. Biotechnol. 60, 155–163.

Kengen, S., Luesink, E., Stams, A., Zehnder, A., 1993. Purification

and characterization of an extremely thermostable b-glucosidase

from the hyperthermophilic archaeon Pyrococccus furiosus. Eur. J.

Biochem. 213, 305–312.

Kenji, S., Akira, Y., Hajime, K., Mamoru, W., Mitsuaki, M., 1994.

Purification and characterization of three thermostable b- N -

acetylhexoseaminidase of Bacillus stearothermophilus CH-4 isolated

from chitin-containing compost. Appl. Environ. Microbiol. 60,

2911–2915.

Kenji, S., Akira, Y., Hajime, K., Mamoru, W., Mitsuaki, M., 1998.

Purification and characterization of three thermostable endoch-

itinases of a noble Bacillus strain, MH-1, isolated from chitin-

containing compost. Appl. Environ. Microbiol. 64, 3397–3402.

Kim, H., Sung, M., Kim, M., Oh, T., 1994. Occurrence of thermo-

stable lipase in thermophilic Bacillus sp. Strain 398. Biosci. Biotech.

Biochem. 58, 961–962.

Kim, C., Nashiru, O., Ko, J., 1996. Purification and biochemical

characterization of pullulanase type I from Thermus caldophilus

Gk-24. FEMS Microbiol. Lett. 138, 147–152.

Kim, I., Kim, H., Lee, K., Chung, S., Ko, S., 2002a. Lipase-catalyzed

acidolysis of perilla oil with caprylic acid to produce structured

lipids. JAOCS 79, 363–367.




Kim, Y., Bae, J., Oh, B., Lee, W., Choi, J., 2002b. Enhancement of

proteolytic enzyme activity excreted from Bacillus stearothermo-

philus for a thermophilic aerobic digestion process. Biores. Tech-

nol. 82, 57–164.

Klaus, D., 1998. An enzyme process for the physical refining of seed

oils. Chem. Eng. Technol. 21, 278–281.

Klibanov, A., 1983. Immobilized enzymes and cells as practical

catalysts. Science 219, 722–727.

Klingberg, M., Hashwa, F., Antrakikian, G., 1991. Properties of extremely thermostable proteases from anaerobic hyperthermo-

philic bacteria. Appl. Microbiol. Biotechnol. 34, 715–719.

Knor, D., 1987. Food biotechnology. In: Tannenbaum, S.R., Walstra,

P. (Eds.), Food Science and Technology. Dekker, New York.

Kocabiyik, S., Erdem, B., 2002. Intracellular alkaline proteases

produced by thermoacidophiles: Detection of protease heteroge-

neity by gelatin zymography and polymerase chain reaction (PCR).

Biores. Technol. 84, 29–33.

Koch, R., Zabalowski, P., Antrakianin, G., 1987. Highly active and

thermostable amylases and pullulanases from various anaerobic

thermophiles. Appl. Microbiol. Biotechnol. 27, 192–198.

Koch, R., Spreinat, K., Lemke, K., Antranikan, G., 1991. Purification

and Properties of a hyperthermoactive a-amylase from the

archaeobacterium Pyrococcus woesei . Arch. Microbiol. 155, 572–

578.Kohilu, U., Nigam, P., Singh, D., Chaudhary, K., 2001. Thermostable,

alkaliphilic and cellulase free xylanases production by Thermoac-

tinomyces thalophilus subgroup C. Enzyme Microb. Technol. 28,

606–610.

Konishi, Y., Amemura, A., Tanabe, S., Harada, T., 1979. Immuno-

logical study of pullulanase from Klebsiela strains and the

occurrence of this enzyme in the Enterobacteriaceae. Int. J. Syst.

Bacteriol. 29, 13–18.

Krahe, M., Antranikian, G., Markel, H., 1996. Fermentation of

extremophilic microorganisms. FEMS Microbiol. Rev. 18, 271–

285.

Krishnan, T., Chandra, A., 1983. Purification and characterization of

a-amylase from B. licheniformis. Appl. Environ.Microbiol. 46, 430–

437.

Kubo, M., Imanaka, T., 1988. Cloning and nucleotide sequence of thehighly thermostable neutral protease gene from Bacillus stearo-

thermophilus. J. Gen. Microbiol. 134, 1883–1892.

Kulkurani, N., Gadre, R., 1999. A novel alkaline, thermostable,

protease free lipase from Pseudomonas sp. Biotechnol. Lett. 21,

897–899.

Kumar, S., Nussinov, R., 2001. How do thermophilic proteins deal

with heat? A review. Cell. Mol. Life Sci. 58, 1216–1233.

Kumar, H.D., Swati, S., 2001. Modern Concepts of Microbiology,

second revised ed. Vikas Publishing House Pvt. Ltd., New Delhi.

Kwak, Y., Akeba, T., Kudo, T., 1998. Purification and characteriza-

tion of enzyme from hyperthermophilic archaeon Thermococcus

profundus, which hydrolyses both a-1-4 and a-1-6 glucosidic

linkages. J. Ferment. Bioeng. 86, 363–367.

Laderman, K., Davis, B., Krutzsch, H., Lewis, M., Griko, Y.,

Privalov, P., Anfinsen, C., 1993a. The purification and character-

ization of an extremely thermostable a-amylase from the hyper-

thermophilic archaebacterium Pyrococcus furiosus. J. Biol. Chem.

268, 24394–24401.

Landerman, K., Asada, K., Uemori, T., Mukai, H., Taguchi, Y.,

Kato, I., Anfinsen, C., 1993b. Alpha amylase from the hyper-

thermophilic archaebacterium Pyrococcus furious. Cloning and

sequencing of the gene and expression in E. coli . J. Biol. Chem. 268,

24402–24407.

Lasa, I., Berenguer, J., 1993. Thermophilic enzymes and their

biotechnological potential. Microbiologia 9, 77–89.

Lee, S., Rhee, J., 1993. Production and partial purification of a lipase

from Pseudomonas putida 3 SK. Enzyme Microbiol. Technol. 15,

617–623.

Lee, Y., Lowe, S., Zeikus, J., 1998. Molecular biology and biochem-

istry of xylan degradation by thermoanaerobes. In: Kalegoris, E.,

Christakopoulos, D., Kekos, D., Macris, B. (Eds.), Studies on the

solid-state production of thermostable endoxylanases from Ther-

moascus aurantiacus: characterization of two isoenzymes. J.

Biotechnol. 60, 155–163.

Leuschner, C., Antranikan, G., 1995. Heat stable enzymes from

extremely thermophilic and hyperthermophilic microorganisms.

World J. Microbiol. Biotechnol. 11, 95–114.Ling, L., Keohavong, P., Dias, C., Thilly, W., 1991. Optimization

of the polymerase chain reaction with regard to fidelty: modified

T7, Taq, and Vent DNA polymerases. PCR Meth. Appl. 1, 63–

69.

Linko, Y., Lamsa, M., Huhatala, A., Linko, P., 1994. Lipase catalyzed

transesterification of rapeseed oil snf 2-ethyl-1-hexanol. JAOCS 71,

1411–1414.

Longley, M., Bennet, S., Mosbaugh, D., 1990. Characterization of the

50 –300 exonuclease associated with Thermus aquqticus DNA poly-

merase. Nucleic Acids Res. 18, 7317–7322.

Lopez, G., 1999. DNA supercoiling and temperature adaptation: a

clue to early diversification of life. J. Mol. Evol. 46, 439–452.

Luisa, M.R., Schmidt, C., Wahl, S., Sprauer, A., Schmid, R., 1997.

Thermoalkalophilic lipase of Bacillus thermocatenulatus. Large

scale production, purification and properties: aggregation behaviorand its effect on activity. J. Biotechnol. 56, 89–102.

Lundberg, K., Shoemaker, D., Adams, M., Short, J., Sorge, J.,

Marthur, E., 1991. High-fidelty amplification using a thermostable

polymerase isolated from Pyrococcus furiosus. Gene 108, 1–6.

Majeti, N., Kumar, R., 2000. A review of chitin and chitosan

applications. React. Funct. Polym. 46, 1–27.

Mamo, G., Gessese, A., 1999. A highly thermostable amylase from a

newly isolated thermophilic Bacillus sp. WN11. J. Appl. Microbiol.

86, 557–560.

Manchini, P., Foretina, M., 1998. Production in sea–water of

thermostable alkaline protease by a halotolerant strain of Bacillus

licheniformis. Biotechnol. Lett. 20, 565–568.

Manchini, P., Foretina, M., Parini, C., 1988. Thermostable alkaline

protease produced by Bacillus thermorubber: a new species of

Bacillus. Appl. Microbiol. Biotechnol. 28, 409–413.Manco, G., Adinolf, E., Pisani, F., Ottolina, G., Carea, G., Rossi, M.,

1998. Overexpression and properties of a new thermophilc and

thermostable esterase from Bacillus acidocaldarius with sequence

similarity to hormone-sensitive lipase subfamily. Biochem. J. 332,

203–212.

Mao, W., Pan, R., Freedman, D., 1992. High production of alkaline

protease by Bacillus licheniformis in fed-batch fermentation using

synthetic medium. J. Ind. Microbiol. 11, 1–6.

Mariella, R., 2001. Development of a battery-powered, hand-held,

real-time PCR instrument. Proc. SPIE––Int. Soc. Opt. Eng. 4265,

58–64.

Marjaana, R., Indra, M., Birgitte, A., Liisa, V., 1994. Application of

thermostable xylanase of Dictyoglomus sp. in enzymatic treatment

of kraft pulps. Appl. Microbiol. Biotechnol. 41, 130–139.

Matsui, I., Sakai, Y., Matsui, E., Kikuchi, H., Kawarabayasi, Y.,

Honda, K., 2000. Novel substrate specificity of a membrane-bound

b-glycosidase from the hyperthermophilic archaeon Pyrococcus

horikoshi . FEBS Lett. 467, 195–200.

Mawadza, C., Hatti-Kaul, R., Zvauya, R., Mattiasson, B., 2000.

Purification and characterization of cellulases produced by two

Bacillus strains. J. Biotechnol. 83, 177–187.

Mayer, J., Lupas, A., Kellerman, J., Eckerskorn, C., Baumeister, W.,

Peters, J., 1996. A hyperthermostable protease of the subtilisin

family bound to the surface of the layer of the archaeon

Staphylothermus marinus. Curr. Biol. 6, 739–749.

Mead, D., McClary, J., Luckey, J., Kostichka, A., Witney, F., Smith,

L., 1991. Bst DNA polymerase permits rapid sequence analysis

from nanogram amounts of template. Biotechniques 11, 76–78.




Medda, S., Chandra, K., 1980. New strains of B. licheniformis and B.

coagulans producing thermostable a-amylases active at alkaline

pH. J. Appl. Bacteriol. 48, 47–58.

Min Ha, Y., Lee, D., Yoon, J., Park, Y., Kim, Y., 2001. Rapid and

simple purification of a novel extracellular b-amylase from Bacillus

sp. Biotechnol. Lett. 23, 1435–1438.

Miura, S., Kubota, N., Kawakita, H., Saito, K., Sugita, K., Watanabe,

K., Sugo, T., 2002. High-throughput hydrolysis of starch during

permeation across alpha-amylase-immobilized porous hollow-fibermembranes. Radiat. Phys. Chem. 63, 143–149.

Morgan, F., Priest, F., 1981. Characterization of thermostable a-

amylases from B. licheniformis. J. Appl. Bacteriol. 50, 107–114.

Morris, D., Gibbs, M., Chin, C., Koh, H., Wong, K., Allison, R.,

Nelson, P., Bergquist, P., 1998. Cloning of the xynB gene form

Dictyoglomus thermophilum Rt46B.1 and action of the gene

product on kraft pulp. Appl. Environ. Microb. 64, 1759–1765.

Mozersky, S., Marmer, W., Dale, A.O., 2002. Vigorous proteolysis:

Relining in the presence of an alkaline protease and bating (Post-

Liming) with an extremophile protease. JALCA 97, 150–155.

Mozhaev, V., 1993. Mechanism-based strategies for protein thermo-

stabilization. Trends Biotechnol. 11, 88–95.

Mullis, K., Faloona, F., 1987. Specific synthesis of DNA in vitro via a

polymerase-catalyzed chain reaction. Meth. Enzymol. 155, 335–

350.Mullis, K., Faloona, F., Saiki, R., Horn, G., Erlich, H., 1986. Specific

enzymatic amplification of DNA in vitro: the polymerase chain

reaction. Cold Spring Harbor Symp. Quant. Biol. 51, 263–273.

Myers, T., Gelfand, D., 1991. Reverse transcription and DNA

amplification by a Thermus thermophilus DNA polymerase.

Biochemistry 30, 7661–7665.

Nagao, T., Shimada, Y., Sugihara, A., Murata, A., Komemushi, S.,

Tominaga, Y., 2001. Use of thermostable Fusarium heterosporum

lipase for production of structured lipid containing oleic and

palmitic acids in organic solvent-free system. JAOCS 78, 167–172.

Nakamura, N., Watanabe, K., Horikoshi, K., 1975. Purification and

some properties of alkaline pullulanases from a strain of Bacillus.

Biochem. Biophys. Acta 379, 188–193.

Nakamura, H., Kubota, H., Kono, T., Isogai, A., Onabe, F., 2001.

Modification of pulp properties by cellulase treatment and appli-cation of cellulase to wastepaper deinking and mechanical pulp

refining. In: 68th Pulp and Paper Research Conference. Proceed-

ings of the Pulp and Paper Research Conference, 18–19/06, Japan,

pp. 2–5.

Nawani, N., Dosanjh, N., Kaur, J., 1998. A novel thermostable lipase

from a thermophilic Bacillus sp.: characterization and esterification

studies. Biotecnol. Lett. 20, 997–1000.

Nazan, A., Zumrut, B., 2000. A vicel adsorbable endoglucanase

production by the thermophilic fungus Scytalidium thermophilum

type culture Torula thermophila. Enzyme Microb. Technol. 27,

560–569.

Niehaus, F., Bertoldo, C., Kahler, M., Antranikian, G., 1999.

Extremophiles as a source of novel enzymes for industrial

applications. Appl. Microbiol. Biotechnol. 51, 711–729.

Nipkow, A., Shen, G., Zeikus, J., 1989. Continuos production of

thermostable b-amylase with Clostridium thermosulfurogenes: effect

of culture conditions and metabolite levels on enzyme synthesis and

activity. Appl. Environ. Microbiol. 55, 689–694.

Norman, B., 1979. The application of polysaccharide degrading

enzymes in starch industry. In: Berkley, R. et al. (Eds.), Microbial

Polysaccharides. Academic Press, New York, pp. 339–376.

Nucci, R., Moracci, M., Vaccaro, C., Vespa, N., Rossi, M., 1993.

Exoglucosidase activity and substrate specificity of the b-glucosi-

dase isolated from the extreme thermophilic Sulfolobus solfataricus.

Biotechnol. Appl. Biochem. 17, 239–250.

Nwe, N., Stevens, W., 2002. Production of fungal chitosan by solid

substrate fermentation followed by enzymatic extraction. Biotech-

nol. Lett. 24, 131–134.

Pandey, A., Benjamin, S., Soccol, C., Nigam, P., Krieger, N., Soccol,

V., 1999. The realm of microbial lipases in biotechnology: a review.

Biotechnol. Appl. Biochem. 29, 119–131.

Pantazaki, A., Prista, A., Kyriakidis, D., 2002. Biotechnologically

relevant enzymes from Thermus thermophilus. Appl. Microbiol.


Park, Y., Kang, S., Lee, J., Hong, S., Kim, S., 2002. Xylanase

production in solid state fermentation by Aspergillus niger mutant

using statistical experimental designs. Appl. Microbiol. Biotechnol.58, 761–766.

Paula, S., Alexandra, M., Jose, C., Maria, R., Maria, C., 2002. Rapid

detection of thermostable cellulase free xylanases by a strain of

Bacillus subtilis and its properties. Enzyme Microbiol. Technol. 30,

924–933.

Pawinee, K., Wipapat, K., 1996. Thermostable metalloproteases

excreted by gram-negative thermophillic bacterium. In: Poster

Presentation: 10th International Biotechnology symposium. Syd-

ney, Australia.

Perler, F., Kumar, S., Kong, H., 1996. Thermostable DNA polyme-

rases. Adv. Protein Chem. 48, 377–435.

Peter, W., Alexandra, T., Klaus, D., 2001. Conversions of glycerol to

1,3-propandiol by a newly isolated thermophilic strain. Biotechnol.

Lett. 23, 463–466.

Piller, K., Daniel, R., Petach, H., 1996. Properties and stabilization of an extracellular a-glucosidase from the extremely thermophilic

archae-bacteria Thermococcus strain AN1: enzyme activity at 130

°C. Biochem. Biophys. Acta 1292, 197–205.

Plant, A., Morgan, H., Daniel, R., 1986. A highly stable pullulanase

from Thermus aquaticus. Enzyme Microb. Technol. 8, 668–672.

Poonam, N., Dalel, S., 1995. Enzyme and microbial systems involved

in starch processing. Enzyme Microb. Technol. 17, 770–778.

Rahman, R., Razak, C., Ampon, K., Basri, M., Yunus, W., Salleh, A.,

1994. Purification and characterisation of a heat stable protease

from Bacillus stearothermophilus F1. Appl. Microbiol. Biotechnol.

40, 822–827.

Rakshit, S.K., 1998. Utilization of starch industry wastes. In: Martin,

A. (Ed.), Bioconversion of Waste Materials to Industrial Products,

second ed. Blackie Academic and Professional, London.

Rama, M., Swamy, M., Seenayya, G., 1998. Purification and charac-terization of thermostable b-amylase and pullulanase from high

yielding Clostridium thermosulfurogenes SV2. World J. Microbiol.


Rama, M.,Gopal,R., Seenayya, G.,1999.Enhanced production of ther-

mostable b-amylase and pullulanase in the presence of surfactants

by Clostridium thermosulfurogenes SV2. Proc. Biochem. 34, 87–92.

Ramkrishna, S., Samir, K., Chakrabarty, S., 1993. Immobilization of

a-amylase from Mycelophthora thermophila D-14 (ATCC 48104).

Enzyme Microbiol. Technol. 15, 801–804.

Rao, M., Tankasale, A., Ghatge, M., Desphande, V., 1998. Molecular

and biotechnological aspects of microbial proteases. Microbiol.

Mol. Biol. Rev. 62, 597–634.

Rathi, P., Sapna, B., Sexena, R., Gupta, R., 2000. A hyperthermo-

stable, alkaline lipase from Pseudomonas sp. with the property of

thermal activation. Biotechnol. Lett. 22, 495–498.

Razak, C., Samad, M., Basri, M., Yunus, W., Ampon, K., Salleh, A.,

1993. Thermostable extracellular protease by B. stearothermophi-

lus. World J. Microbiol. Biotechnol. 10, 260–263.

Razak, C., Rahman, R., Salleh, A., Yunus, W., Ampon, K., Basri, M.,

1995. Production of a thermostable protease from a new high pH

isolate of Bacillus stearothermophilus. J. Biosci. 6, 94–100.

Razak, C., Tang, S., Basri, M., Salleh, A., 1997. Prelimenary study on

the production of extracellular protease from a newly isolated

Bacillus sp. (no. 1) and the physical factors affecting its production,

Pertanika. J. Sci. Technol. 5, 169–177.

Rina, R., Geeta, N., 1996. Microbial b-amylase: Biosynthesis, char-

acterization and industrial applications. Crit. Rev. in Microbiol.

22, 181–199.




Roberts, G., 1992. Physical structures. In: Roberts, G.A.F. (Ed.),

Chitin Chemistry, first ed. Macmillan, London, pp. 20–35.

Rolfsmeir, M., Blum, P., 1995. Purification and characterization of a

maltose from the extremely thermophilic crenarchaeoteSulfolobus

solfataricus. J. Bacteriol. 177, 482–485.

Rothschild, L., Manicineli, R., 2001. Life in extreme environments.

Nature 409, 1092–1101.

Rudiger, A., Jorgensen, P., Antranikan, G., 1995. Isolation and

characterization of a heat stable pullulanase from the hyperthermo-philic Archeon Pyrococcus woesei after cloning and expression of

its gene in Escherichia coli . Appl. Environ. Microbiol. 61, 567–575.

Ruthersmith, L., Daniel, R., 1991. Thermostable cellobiohydrolase

from the thermophilic eubacterium Thermotoga sp. Strain FjSS3-

B.1. Purification and properties. Biochem. J. 277, 887–890.

Ruthersmith, L., Daniel, R., 1992. Cellulytic and hemicellulytic

enzymes functional above 100 °C. Ann. NY Acad. Sci. 672, 137–

141.

Saboto, D., Nucci, R., Rossi, M., Gryczynski, I., Gryczyniski, Z.,

Lakowicz, J., 1999. The b-glycosidase from the hyperthermophilic

archaeon Sulfolobus solfataricus: enzyme activity and conforma-

tional dynamics at temperatures above 100 °C. Biophys. Chem. 81,

23–31.

Saha, B., Mathupala, S., Zeikus, J., 1988. Purification and characteri-

zation of a highly thermostable novel pullulanase from Clostridiumthermohydrosulfuricum . Biochem. J. 252, 343–348.

Saiki, R., Gelfand, D., Stoffe, S., Scharf, S., Higuchi, R., Horn, G.,

Mullis, K., Erlich, H., 1988. Primer directed enzymatic amplifica-

tion of DNA with thermostable DNA polymerase. Science 239,

487–491.

Salleh, A.B., Basri, M., Razak, C., 1977. The effect of temperature on

the protease from Bacillus stearothermophilus strain F1. Mal. J.

Biochem. Mol. Biol. 2, 37–41.

Sarikaya, E., Higassa, T., Adachi, M., Mikami, B., 2000. Comparison

of degradation abilities of a- and b-amylases on raw starch

granules. Proc. Biochem. 35, 711–715.

Satosi, H., Seigo, O., Kenji, T., Kazuhisa, K., Tetsuo, K., Toshiaki,

K., Hitosi, K., 2001. Chemo-enzymatic synthesis of 3-(2-naphtyl)-

LL-alanine by an amino transferase from the extreme thermophiles

Thermococcus profoundus. Biotechnol. Lett. 23, 589–591.Saul, D., Williams, L., Reeves, R., Gibbs, M., Bergquist, P., 1995.

Sequence and expression of a xylanase gene from the hyperther-

mophile Thermotoga sp. strain FjSS3-B1 and characterization of

the recombinant enzyme and its activity on Kraft pulp. Appl.

Environ. Microb. 61, 4110–4113.

Schmidt, C., Sztajer, H., Stocklein, W., Menge, U., Schimid, R., 1994.

Screening, purification and properties of a thermophilic lipase from

Bacillus thermocatelnulatus. Biochem. Biophys. Acta 1214, 43–53.

Sebastien, Z., Jean-Luk, R., Dider, F., Yannik, G., Joseph, B.,

Jacques, D., 2001. Characterization of highly thermostable alkaline

phosphatase from the Euryarchaeon Pyrococcus abyssi . Appl.

Environ. Microbiol. 67, 4504–4511.

Sharma, R., Soni, S., Vohra, R., Gupta, L., Gupta, J., 2002.

Purification and characterization of a thermostable alkaline lipase

from a new thermophilic Bacillus sp. RSJ-1. Proc. Biochem. 37,

1075–1084.

Shaw, J., Pang, L., Chen, S., Chen, I., 1995. Purification and properties

of an extracellular a-amylase from Thermus sp. Bot. Bull. Acad.

Sci. 36, 195–200.

Shen, G., Saha, B., Lee, Y., Bhatnagar, L., Zeikus, J., 1988.

Purification and characterization of a novel thermostable b-

amylase from Clostridium thermosulfurogenes. Biochem. J. 254,

835–840.

Shimada, Y., Watanabe, Y., Samukawa, T., Sugihara, A., Noda, H.,

Fukuda, H., Tominaga, Y., 1999. Conversion of vegetable oil to

biodiesel using immobilized Candida antarctica lipase. JAOCS 76,

789–793.

Simpson, H., Haufler, U., Daniel, K., 1991. An extremely thermostable

xylanase from the thermophilic eubacterium Thermotoga. Biochem.

J. 277, 177–185.

Spano, L., Medeiros, J., Mandels, M., 1975. In: Enzymatic Hydrolysis

of Cellulosic Waste to Glucose (Pollution Abatement Div., Food

Svcs. Lab, US Army Natick Labs), Natick, MA.

Spindler, K., Spindler, B.M., Londershausen, M., 1990. Chitin

metabolism: a target for drugs against parasites. Parasitol. Res.

76, 283–288.Stamford, T., Stamford, N., Coelho, L., Araujo, J., 2001. Production

and characterization of thermostable. Endophyte of yam bean a-

amylase from Nocardiopsis sp. Biores. Technol. 76, 137–141.

Stetter, K., 1999. Extremophiles and their adaptation to hot environ-

ments. FEBS Lett. 452, 22–25.

Sunna, A., Moracci, M., Rossi, M., Antranikian, G., 1996a. Glycosyl

hydrolases from hyperthermophiles. Extremophiles 1, 2–13.

Suntornsuk, W., Pochanakanich, P., Suntornsuk, L., 2002. Fungal

chitosan production on food processing by-products. Proc. Bio-

chem. 37, 727–729.

Suzuki, Y., Chishoro, M., 1983. Production of extracellular thermo-

stable pullulanase by an amylolytic obligatory thermophilic soil

bacterium B. stearothermophilus. Eur. J. Appl. Biotechnol. 17, 24–

29.

Swampy, M., Sai Ram, M., Seenayya, G., 1994. b-amylase fromClostridium thermocellum SS8––a thermophilic, anaerobic, cellu-

lytic bacterium. Lett. Appl. Microbiol. 18, 301–304.

Swamy, M., Seenayya, G., 1996. Thermostable pullulanase and a-

amylase activity from Clostridium thermosulfurogenes SV9-optimi-

zation of culture conditions from enzymes production. Proc.

Biochem. 31, 157–162.

Swaroopa, D., Krishna, N., 2000. Production of thermostable cellu-

lase-free xylanases by Clostridium absonum CFR-702. Proc.

Biochem. 36, 355–362.

Takami, H., Akiba, T., Horikoshi, K., 1989. Production of extremely

thermostable alkaline protease from Bacillus sp. Appl. Microbiol.


Takasaki, Y., 1976a. Production and utilization of b-amylase and

pullulanase from Bacillus cereus var. mycoides. Agri. Biol. Chem.

40, 1515–1522.Takasaki, Y., 1976b. Purification and enzyme properties of b-amylase

and pullulanase from Bacillus cereus var. mycoides. Agri. Biol.

Chem. 40, 1523–1530.

Takayanagi, T., Ajisaka, K., Takiguchi, Y., Shimahara, K., 1991.

Isolation and characterization of thermostable chitinases from

Bacillus lichiniformis X_7u. Biochem. Biophys. Acta 1078, 404–410.

Takizawa, N., Muroorka, Y., 1985. Cloning of the pullulanase gene in

E. coli and Klebsiella aerogenes. Appl. Environ. Microbiol. 49, 294–

298.

Tan, K., Gill, C., 1985. Effect of culture conditions on batch growth of

Pseudomonas fluorescens on olive oil. Appl. Microbiol. Biotechnol.

23, 27–32.

Traore, M.K., Buschle-Diller, G., 2000. Environmentally friendly

scouring processes. Textile Chem. Colorist Am. Dye Stuff Report.

32, 40–43.

Tsujibo, H., Endo, H., Miyamoto, K., Inamori, Y., 1995. Expression

in Escherichia coli of a gene encoding a thermostable chitinase from

Streptomyces thermoviolaceus OPC-520. Biosci. Biotechnol. Bio-

chem. 59, 145–146.

Underkofler, L., 1976. Microbial enzymes. In: Miller, B., Litsky, W.

(Eds.), Industrial Microbiology. McGraw-Hill, New York.

Van der Maarel, M., Van der Veen, B., Uitdehaag, H., Leemhuis, H.,

Dijkhuizen, L., 2002. Properties and applications of starch-

converting enzymes of the a-amylase family. J. Biotechnol. 94,

137–155.

Van wyk, J., Mogale,A., Seseng, T., 2001. Bioconversion of wastepaper

to sugars by cellulase from Aspergillus niger, Trichoderma viride




and Penicillium funiculosum. J. Solid Waste Technol. Manage. 27,

82–86.

Velikodvorskaya, T., Volkov, I., Vasilenko, V., Zverlov, V., Pirusian,

E., 1997. Purification and some properties of Thermotoga neapol-

itana thermostable xylanase B expressed in E. coli cells. Biochem-

istry 62, 66–70.

Viara, N., Elena, P., Elka, I., 1993. Purification and characterization of

a thermostable alpha-amylase from Bacillus licheniformis. J.

Biotechnol. 28, 277–289.Vihinen, M., Mantsala, P., 1990. Characterization of a thermostable

Bacillus stearothermophilus alpha-amylase. Biotechnol. Appl. Bio-

chem. 12, 427–435.

Vikari, L., Kantelinen, A., Sundquist, J., Linko, M., 1994. Xylanases

in bleaching: from an idea to the industry. FEMS Microbiol. Rev.

13, 335–350.

Volkl, P., Stetter, K., Miller, J., 1995. The sequence of a subtilin-type

protease (aerolysin) from the hyperthermophilic archaeon Pyro-

baculum aerophilum reveals sites important to thermostability.

Protein Sci. 3, 1329–1340.

Vulfson, E., 1994. Industrial applications of lipases. In: Wooley, P.,

Peterson, S.B. (Eds.), Lipases: Their Structure, Biochemistry, and

Application. Cambridge University Press, Cambridge, UK, pp.

271–288.

William, C., Dennis, C., 1988. Food Microbiology, international ed.McGraw-Hill, NY.

Winterhalter, C., Liebl, W., 1995. Two extremely thermostable

xylanases of the hyperthemophilic bacterium. Thermotoga mari-

tema MSB8. Appl. Environ. Microbiol. 61, 1810–1815.

Woese, C., Kandler, O., Wheelis, M., 1990. Towards a natural system

of organisms: Proposal for the domains Archaea, Bacteria, and

Eucara. Proc. Natl. Acad. Sci. USA 87, 4576–4579.

Wojciechowski, P.M., Koziol, A., Noworyta, A., 2001. Iteration model

of starch hydrolysis by amylolytic enzymes. Biotechnol. Bioeng. 75,

530–539.

Wu, X., Jaaskelainen, S., Linko, Y., 1996. An investigation of crude

lipase for hydrolysis, esterification, and transesterification. Enzyme

Microb. Technol. 19, 226–231.

Yan, F., Yong-Goe, J., Kazuhiko, I., Hiroyasu, I., Susumu, A., Tohru,

Y., Hiroshi, N., Shugui, C., Ikuo, M., Yoshitsugu, K., 2000.

Thermophilic phospholipase A2 in the cytosolic fraction from the

archaeon Pyrococcus horikoshii . JAOCS 77, 1075–1084.

Young, M., Dong, G., Jung-Hoon, Y., Yong-Ha, P., Young, J., 2001.

Rapid and simple purification of a novel b-amylase from Bacillussp. Biotechnol. Lett. 23, 1435–1438.

Zeikus, J., Lee, C., Lee, Y., Saha, B., 1998a. Thermostable saccharides:

new sources, uses and biodesigns. In: Kalegoris, E., Christakopo-

ulos, D., Kekos, D., Macris, B. (Eds.), Studies on the Solid-State

Production of Thermostable Endoxylanases from Thermoascus

aurantiacus: Characterization of Two Isoenzymes. J. Biotechnol.

60, 155–163.

Zeikus, J., Vielle, C., Savachenko, A., 1998b. Thermozymes: biotech-

nology and structure-function relationships. Extremophiles 2, 179–

183.

Zhang, J., Greasham, R., 1999. Chemically defined media for

commercial fermentations. Appl. Microbiol. Biotechnol. 51, 407–

421.

Zhou, L., Yeung, K, Yuen, C., 2001. Combined cellulase and wrinkle-

free treatment on cotton fabric. J. Dong Hua University 18, 11– 15.

Zverlov, V., Piotukh, K., Dakhova, O., Velikodvorskaya, G., Borris,

R., 1996. The multidomain xylanase A of hyperthermophilic

bacterium Thermotoga neapolitana is extremely thermoresistant.

Appl. Microbiol. Biotechnol. 45, 245–247.

Zverlov, V., Riedel, K., Bronnenmeier, K., 1998. Properties and gene

structure of a bifunctional cellulytic enzyme (CelA) from the

extreme thermophile Anaerocellum thermophilum with separate

glycosyl hydrolase family 9 and 48 catalytic domains. Microbiology

144, 457–465.


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