enzymes and enzyme kinetics 2012

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Page 1: Enzymes and enzyme kinetics 2012

ENZYMES

Page 2: Enzymes and enzyme kinetics 2012

OBJECTIVESAt the end of the lecture, the student should be able to: 1. Define the following: a. Enzymes b. Apoenzyme c. Coenzyme d. Holoenzyme e. Metalloenzyme f. Regulatory enzyme g. Active site of the enzyme h. Allosteric site of the enzyme i. Substrate 2. Discuss the helpers (cofactors) of enzymes.

Page 3: Enzymes and enzyme kinetics 2012

3. Enumerate the six major classes of enzymes.4. Discuss the characteristics of enzymes. 5. Explain the models of enzyme-substrate complex. 6. Explain enzyme kinetics. a. Factors that affect enzyme activity or

reaction velocity.b. Ways of expressing enzyme activity.

7. Discuss the operation and plots used to illustrate enzyme kinetics.

a. Michaelis-Menten kineticsb. Lineweaver-Burke Double Reciprocal Plotc. Michaelis constant and its significanced. Kinetic order of reactions

OBJECTIVES

Page 4: Enzymes and enzyme kinetics 2012

8. Discuss enzyme inhibition and its effect on reaction velocity. a. Reversible b. Irreversible9. Discuss the different ways of regulating enzyme activity.10. Explain the factors affecting enzyme activity.11. Elucidate uses and clinical application of

enzymes.

OBJECTIVES

Page 5: Enzymes and enzyme kinetics 2012

ENZYMES

Specialized protein catalysts that accelerate chemical

reactions

Page 6: Enzymes and enzyme kinetics 2012

APOENZYME APOENZYME APOENZYME

DEFINITION OF TERMS

Protein part

Cofactor (Nonprotein

part)Coenzyme

Prosthetic group

Metal ion

HOLOENZYME

+ ++

Page 7: Enzymes and enzyme kinetics 2012

ENZYME COFACTORS

A. Coenzyme EnzymeChemical Groups

Transferred

VitaminPrecursor

Thiamine Pyrophosphate

(TPP)

Pyruvate dehydrogenase, Isocitrate dehydrogenase, α-ketoglutarate dehydrogenase, Transketolase, α-Ketoacid dehydrogenase

Aldehydes Thiamine (Vit B1)

Flavin Adenine Dinucleotide

(FAD)

Succinate dehydrogenase, α-Ketoglutarate dehydrogenase, Pyruvate dehydrogenase, Nitric oxide synthase

ElectronsRiboflavin

(Vit B2)

Nicotinamide Adenine

Dinucleotide (NAD)

Lactate dehydrogenase; Other dehydrogenases

Hydride ion(:H-)

Nicotinic acid(Niacin; B3)

Pyridoxal Phosphate (PLP)

Glycogen phosphorylase, γ-ALA synthase, Histidine decardoxylase, Alanine aminotransferase

Amino groupsPyridoxine

(Vit B6)

Lipoate Pyruvate dehydrogenaseα-Ketoglutarate dehydrogenase

Electrons and acyl groups

Not required in diet

Page 8: Enzymes and enzyme kinetics 2012

ENZYME COFACTORS

A. Coenzyme EnzymeChemical Groups

Transferred

Vitamin Precursor

Coenzyme A(CoASH)

Acetyl CoA carboxylase Acyl groups

Pantothenic acid & other compounds

Biocytin

Pyruvate carboxylase, Acetyl CoA carboxylase, Propionyl CoA carboxylase

CO2

Biotin

5’-deoxycobalamin

Methylmalonyl mutase

H atoms and alkyl groups

Vit B12

TetrahydrofolalateThmidylate synthase

One-carbon groups

Folic acid

Page 9: Enzymes and enzyme kinetics 2012

CLASSES OF COENZYMES

CLASS EXAMPLES

Activation-Transfer Coenzymes

TPPCoenzyme A

BiotinPLP

Oxidation-ReductionCoenzymes

NAD+

FADVit E, Vit C

Mark’s Medical Biochemistry, 3rd ed.

Page 10: Enzymes and enzyme kinetics 2012

ENZYME COFACTORS: COENZYME A

C-CH2-CH2-N-C-C—C-CH2O

OII

OII

IOH

H

IIO

I CH3

CH3

I

I NH

I CH2

I CH2

I SH

O = P – O-

IOI

O = P –O-

IO

NH2

NN

N N

OI

O = P – O-

IO-

O

OH

Pantothenic acid

1. Pantothenic acid-derived, co-factor of several enzymes like acetyl CoA carboxylase.

2. Takes part in reactions of the CAC, FA synthesis and oxidation, acylations and cholesterol synthesis.

H

H

HH H

Active sulfhydryl

group that form thioesters with

acyl groups

Page 11: Enzymes and enzyme kinetics 2012

COO-

| CH2

| CH2

| C = O |

S ~ CoA

COOH | CH2

| CH2

| C = O | COO-

α-Ketoglutarate (C5) Succinyl CoA (C4)

NAD+ NADH + H+

CO2CoASH

α-Ketoglutarate Dehydrogenase Complex

Coenzymes:1. TPP2. Lipoic Acid3. FAD4. NAD+

5. CoASH

ΔG0 = - 8.0 kcal

ENZYME COFACTORS: COENZYME A

Pyruvate decarboxylaseDihydrolipoyl transacetylaseDihydrolipoyl dehydrogenase

Page 12: Enzymes and enzyme kinetics 2012

COO-

|C=O|CH3

S ~ CoA |C=O |CH3

NAD+NADH+ +

H+

CoASH

Pyruvate Dehydrogenase Complex

TPP LipoateFAD

Pyruvate decarboxylaseDihydrolipoyl transacetylaseDihydrolipoyl dehydrogenase

Pyruvate(C3)

Acetyl CoA(C2)

CO2

ΔG0 = - 8.0 kcal/mole

ENZYME COFACTORS: COENZYME A

Page 13: Enzymes and enzyme kinetics 2012

ENZYME COFACTORS: NAD & NADP

OHOH

O II

C – NH2

NIR

O

-O

N

N

NH2

N

N

O

H

O – CH2

HH

O

H

OH

– CH2

HH

OH

O

-O

P

O

O

-O

P

OII

C – NH2

H

N

NADP+ contains a Pon this 2’-hydroxyl

H

H

Nic

oti

na

mid

e a

den

ine

din

uc

leo

tid

e (

NA

D+)

Adeninering

Ribose ring

+

COO-

I

H-O-C-H I

CH3

Lactate

Dissociates as H+

COO-

I C=O + H+

I CH3

Pyruvate

1

Nicotinamidering

CC

Ketogroup

1

3

2

Functionalgroup

2

1

1’

2’

349

87

65

AMP provides additionalbinding interactions that induce conformational changes in the enzyme

AM

P

Lactatedehydrogenase

3’2’

2’

3’

5’

4’

Page 14: Enzymes and enzyme kinetics 2012

ENZYME COFACTOR: NAD+

COO-

| HO - C – H | CH3

COO-

I C = O

I CH3

Lactate dehydrogenase

L-Lactate PyruvateNAD+ NADH+ + H+

Page 15: Enzymes and enzyme kinetics 2012

H

H

R

H-N N-H

S

IIIIIIIIIIIIIIIIIIIIIIIIIIIIIIH

O

IIIIIIIIIIIIIII

II

H

COENZYME: BIOTIN

CO2

attachment site

NH

NH

CIIO

H

H-N N-H

S

IIIIIIIIIIIIIIIIIIIIIIIIIH

O

IIIIIIIIIII

II

= O

Biotin

Protein portion of enzyme:Acetyl CoA carboxylasePropionyl CoA carboxylasePyruvate carboxylase

Lysyl residue

Biotin

1. One of the B complex vitamins.

2. A cofactor of such enzymes as acetyl

CoA carboxylase, propionyl CoA

carboxylase and pyruvate carboxylase.

3. It is a carrier of activated CO2, hence involved in

carboxylation reactions.

OII

Page 16: Enzymes and enzyme kinetics 2012

COENZYME: BIOTIN

Pyruvate

Gluconeogenesis

Oxaloacetate

ATP ADP + Pi

Pyruvate carboxylase

CO2BIOTIN

COO-

I C =O

I CH3

COO-

|C= O|CH2

|COO-

Page 17: Enzymes and enzyme kinetics 2012

COENZYME:THIAMINE PYROPHOSPHATE

H

Thiamine Pyrophosphate(TPP; Vit B1-derived)

N

H3C

NN NC H

HS

C-C-OHH

AMP

CN

NH2

CH

CNO-

IIO

OII

CH2

H3C

NH2

S

C

C

C

N

H

CH3

- CH2 –CH2 – O – P –O – P – O-

O-

Dissociable proton

Reactivecarbon atom:

carrier of aldehyde groups

Coenzyme binding

site

Page 18: Enzymes and enzyme kinetics 2012

Succinate dehydrogenase

(C6) OXALOACETATE CITRATE (C6)

a – KETOGLUTARATE (C5)

SUCCINYL CoA (C4)

ISOCITRATE (C6)

(C4) FUMARATE

(C4) SUCCINATE

(C4) MALATENAD

NADH + H+

FADH2

FAD

H2O

NAD

NADH + H+

CO2

H2O

H2O

Malate dehydrogenase

Isocitrate dehydrogenase

Succinyl CoA thiokinase

PYRUVATE

ACETYL-CoA

Citrate synthase

NAD

NADH + H+

CO2

Pyruvate

dehydrogenase

(ATP)

GTP ADP + Pi

CoAMg+

NAD

NADH + H+

CO2

Aconitase

Fumarase

a - ketoglutarate dehydrogenase

(C3)

(C2)

ETC

H2O O2

ATPOxid.

Phospho

Role of TPP in the Oxidative Decarbo-

xylation Reactions of CAC

TPP

TPP

TPP

Page 19: Enzymes and enzyme kinetics 2012

H3C

NH2

O

H3C

H3C

NHN

NN O

CH2

I H-C-OH

I H-C-OH

I H-C-OH

I CH2-O-P PO O-CH2

OII

IO

P

NH2

N

N

N

N

O

OH OH

OII

COENZYME: FAD

Flavin Adenine Dinucleotide (FAD)

A riboflavin (Vit B3)-derived coenzyme of several dehydrogenases involved in

oxidation-reduction reactions.

Isoalloxacin ring

Ribitol

From ATP

Page 20: Enzymes and enzyme kinetics 2012

COOH |

CH2

| CH2

| COOH

H -COOH | H – C ||

C – H |

H -COOH

Fumarate (C4)Succinate (C4)FAD FADH2

Succinate dehydrogenase

ROLE OF FAD IN THE CITRIC ACID CYCLE

COENZYME: FAD

Page 21: Enzymes and enzyme kinetics 2012

ROLE OF FAD AND FMN IN NITRIC OXIDE (NO) SYNTHESIS

Arginine

NO +

NADPH2

+ O2

NADP+

+ H2O

Nitric oxide synthaseFAD, FMN, HemeTetrahydrobiopterin

COENZYME: FAD NH2

I H2N = C I

NH I

CH2

I CH2

I CH2

I H3N – C – H

I COO-

NH2

I C = O

I NH

I CH2

I CH2

I CH2

I H3N – C – H

I COO-

+

Citrulline

+

+

Page 22: Enzymes and enzyme kinetics 2012

-O3PO-CH2

CH3

C HO

N

H

OH

OH

Pyridoxal Phosphate

COENZYME:PLP

A Vit. B6-derived coenzyme involved in carbohydrate, amino acid and neurotransmitter synthesis.

+

Reactive aldehyde groupinvolved in the transfer

of amino groups.

Page 23: Enzymes and enzyme kinetics 2012

ROLE OF PLP IN CARBOHYDRATE METABOLISM: GLYCOGENOLYSIS

O

OH

OH

HO

O-PO3=

O

OH

OH

HOO

OH

OH

HO

O

O

OH

OH

HO

O

O

OH

OH

HO

O

Glycogen chain

Glygogenphosphorylase

Pi

O

OH

OH

HO

O

O

OH

OH

HO

O

OH

OH

HO

O+

OH

OH

OHH

OH

Glucose 1-PRemaining glycogen

PLP

H

H

H

H

H

H H

H

H

H

H

H

COENZYME: PLP

Page 24: Enzymes and enzyme kinetics 2012

ROLE OF PLP AS A COENZYME IN AMINOACID METABOLISM: HEME SYNTHESIS

Glycine +Succinyl CoA

δ-Aminolevulenic acid(ALA)

Heme(Fe protoporphyrin IX)

several reactions

δ-Aminolevulenate synthase PLP

COENZYME: PLP

Page 25: Enzymes and enzyme kinetics 2012

ROLE OF PLP IN AMINO ACID METABOLISM: HISTAMINE SYNTHESIS

H|

CH2 – C – COO-

| NH3

PLP

CH2 – CH2 – NH3

Histidine

HISTAMINE

CO2

Histidinedecarboxylase

COENZYME: PLP

Page 26: Enzymes and enzyme kinetics 2012

NH3+

| CH3 – C – C – COO-

|H

Alanine

COO-

| CH2

| CH2

| H– C – NH3

+

| COO-

Glutamate

ROLE OF PLP IN AMINO ACID METABOLISM:TRANSAMINATION

O||

CH3 – C – COO-

Pyruvate

COO-

| CH2

| CH2

| C = O

| COO-

α-Ketoglutarate

+ +

Alanine aminotransferase

(transaminase)

PLP

COENZYME: PLP

Page 27: Enzymes and enzyme kinetics 2012

Cofactor EnzymeB. Inorganic (Metal ions or iron- sulfur clusters)

Zn+2 Carbonic anhydrase, Alcohol dehydroge- nase, Carboxypeptidases A & B

Cu+2 Cytochrome oxidase

Mn+2 Arginase, Ribonucleotide reductase

Mg+2 Hexokinase, Pyruvate kinase, Glucose 6- phosphatase

Ni+2 Urease

Mo Nitrate reductase

Se Glutathione peroxidase

Mn+2 Superoxide dismutase

K+ Propionyl CoA carboxylase

ENZYME COFACTORS

Page 28: Enzymes and enzyme kinetics 2012

ENZYME COFACTORS: Mg+2

Glucose Glucose 6-PO4

HexokinaseGlucokinase

ATP ADP + Pi

Mg+2

O || C1 - H | H - C2 - OH |OH - C3 - H | H - C4 - OH | H - C5 - OH | H - C6 – OH | H

O || C1 - H | H - C2 - OH |OH - C3 - H | H - C4 - OH | H - C5 - OH | H - C6 - O – P | H

Page 29: Enzymes and enzyme kinetics 2012

GLYCOLYSIS

COO-

| C2 = O | CH3

O || C1 – O-

| C2 – O ~ P |H – C3

| H

Phosphoenol Pyruvate(PEP)

PyruvateADP ATP

Mg+2

K+

Δ G0 = - 6.1 kcal/mole

ENZYME COFACTORS: K+

Pyruvate kinase

Page 30: Enzymes and enzyme kinetics 2012

ENZYME COFACTORS: Zn+

CO2 + H2O H2CO3

Carbonic anhydrase

Zn+2

Page 31: Enzymes and enzyme kinetics 2012

METALLOENZYMES

Enzymes that requirea metal in their

composition

Page 32: Enzymes and enzyme kinetics 2012

SUBSTRATE

The molecule acted upon

by the enzyme

to form a

product

Page 33: Enzymes and enzyme kinetics 2012

Hexokinase/Glucokinase

ATP ADP

Mg+2

Δ G0 = - 4.0 kcal/mole

O || C1 - H | H - C2 - OH |OH - C3 - H | H - C4 - OH | H - C5 - OH | H - C6 – OH | H

Glucose

O || C1 - H | H - C2 - OH |OH - C3 - H | H - C4 - OH | H - C5 - OH |

H - C6 - O – P | H

Glucose 6-Phosphate

ATP AS A CO-SUBSTRATE

Page 34: Enzymes and enzyme kinetics 2012

ACTIVE SITE OF THE ENZYME

Page 35: Enzymes and enzyme kinetics 2012

LYSOZOME: ACTIVE SITE

Page 36: Enzymes and enzyme kinetics 2012

CHYMOTRYPSIN:ACTIVE SITE

His 57 Ser 195

Page 37: Enzymes and enzyme kinetics 2012

ALLOSTERIC SITE

Substrate

Enzyme

Allosteric site

Substrate sites

Page 38: Enzymes and enzyme kinetics 2012

REGULATORY ENZYME

The enzyme that catalyzes therate-limiting or committed

step of a metabolicpathway.

Page 39: Enzymes and enzyme kinetics 2012

REGULATORY ENZYME Phosphofructokinase I

Fructose 6-phosphate

Fructose 1,6-bisphosphate

ATP ADP + Pi

Phosphofructokinase I

Glycolysis

H | H - C1 - OH | C2 = O |OH - C3 - H | H - C4 - OH | H - C5 - OH | H - C6 - O – P | H

H | H - C1 - O - P | C2 = O |OH - C3 - H | H - C4 - OH | H - C5 - OH | H - C6 - O – P | H

AMPF 2,6 bisPO4

ATPCitrateH+

+ -

Page 40: Enzymes and enzyme kinetics 2012

O ||CH3 – C – S – CoA

ACETYL CoA

ATP

ADP + Pi

O O \ || C – CH2 – C – S – CoA //O

MALONYL CoA

Acetyl CoA carboxylase

CO2

(HCO3-)

CitrateInsulinHigh CHOLow FatHigh Prot.

Malonyl CoAPalmitoyl CoAEpinephrineGlucagonHigh FatFasting

H2O

+

-

REGULATORY ENZYMEAcetyl CoA Carboxylase

De Novo Synthesis of Fatty Acids

Page 41: Enzymes and enzyme kinetics 2012

HMG CoA Mevalonate

HMG CoA reductase

NADPH + H+

NADPH

CoA

REGULATORY ENZYMEHMG Coa Reductase

Cholesterol Synthesis

C1OO-

| C2H2

| HO – C3 – CH3

| C4H2

| C5H2OH

O II

-O – C I

CH2

I OH –C – CH2

I C – S – CoA

II O

Insulin, T3Glucocorticoids

Cholesterol GlucagonBile acids StatinsMevalonate

-+

Page 42: Enzymes and enzyme kinetics 2012

Glucose 6-phosphate dehydrogenase

NADP NADPH + H+

Pentose Phosphate Pathway

REGULATORY ENZYMEGlucose 6-Phosphate

Dehydrogenase H |

C1 = O |

H – C2 – OH |

HO – C 3– H |

H – C4 – OH |

H – C 5– OH |

C6H2OPO32-

O ||

C1 |

H – C2 – OH |

HO – C3 – H |

H – C4 – OH | H – C5

|

C6H2OPO32-

Glucose 6-phosphate 6-phosphogluconolactone

Page 43: Enzymes and enzyme kinetics 2012

INTRACELLULAR LOCATION OF SOME IMPORTANT BIOCHEMICAL PATHWAYS

Page 44: Enzymes and enzyme kinetics 2012

ISOENZYME

Different structural forms of an enzyme which catalyze the same chemical reactions → act on the same substrate(s) and produce the same product(s) but exhibit differing degrees of efficiency.

Different isoenzymes are expressed in specific tissues of the body.

Page 45: Enzymes and enzyme kinetics 2012

ISOENZYMES OF LACTATE DEHYDROGENASE

Lactate dehydrogenase (LDH) – catalyzes the reversible conversion of pyruvate to lactate.

Tetramer consisting of 2 subunits: M (found in skeletal muscles and

liver) & H (heart).5 distinct isoenzyme forms (from combination of M & H isozymes).

An increase of H4 in the blood indicates tissue damage as in heart attack.

Enzymes can therefore serve as markers for disease.

Page 46: Enzymes and enzyme kinetics 2012

SIX MAJOR CLASSES OF ENZYMES (IUBMB*, 1964)CLASS EXAMPLE

Oxidoreductases Dehydrogenases, Oxidases, Reductases, Peroxidases, Catalases, Oxygenases, Hydroxylases

Transferases Transaldolase and Transketolase; Acyl, methyl glucosyl, and phosphoryltransferases,Kinases, Phosphomutases, Transaminases

Hydrolases Esterases, Glycosidases, Peptidases, Phosphatases, Thiolases, Phospholipases, Amidases, Deaminases, Ribonucleases

Lyases Decarboxylases, Aldolases, Hydratases, Dehydratases, Synthases, Lyases

Isomerases Epimerases, Isomerases, Mutases, Racemases

Ligases Synthetases, Carboxylases

*International Union of Biochemistry and Molecular Biology; classification is based on the reactions enzymes catalyze; each class is divided into subclasses.

Page 47: Enzymes and enzyme kinetics 2012

OXIDOREDUCTASES

Transfer of electrons and hydrogen atoms from donors (or reductants,

hence oxidized to acceptors (or oxidants, hence reduced).

COO-

| HO – C – H + NAD+

| CH3

L-Lactate

COO-

| C = O + NADH + H+

| CH3

Pyruvate

Lactate dehydrogenase

Page 48: Enzymes and enzyme kinetics 2012

TRANSFERASESTransfer functional groups (like C-, N-, or

P-) from donors to acceptors; utilize 2 substrates to produce 2 products.

COO-

| H3N – C – H + C = O | | CH3 (CH2)2

L-Alanine | COO- α-Ketoglutarate

(keto acid)

COO- COO-

| | C = O + H3N – C – O | | CH3 (CH2)2

Pyruvate | COO-

L-Glutamate (amino acid)

Alaninetransaminase

PLP

(amino acid) (keto acid)substrate

substrate

product

product

Page 49: Enzymes and enzyme kinetics 2012

VI. Reactions: GLYCOLYSIS Kinase - transfers the functional group phosphate from ATP to an acceptor

ATP(donor)

ADP + Pi(product)

Mg+2

Δ G0 = - 4.0 kcal/mole

O || C1 - H | H - C2 - OH |OH - C3 - H | H - C4 - OH | H - C5 - OH | H - C6 – OH | H

Glucose(acceptor)

O || C1 - H | H - C2 - OH |OH - C3 - H | H - C4 - OH | H - C5 - OH | H - C6 - O – P | HGlucose 6-phosphate(product)

TRANSFERASES

HexokinaseGlucokinase

Page 50: Enzymes and enzyme kinetics 2012

OHI

CH-C=C-(CH2)12-CH3H

H0II

R1-C-N-CHI

CH2

H

O

UDP-galact-ose

UDP

Fatty acid

Galactose

Galactosyl transferase

OHI

CH-C=C-(CH2)12-CH3

Ceramide

0II

R1-C-N-CHI

CH2OH

H

H

H

Cerebroside (Galactocerebroside; a glycosphingolipid)

H

O

H OH

HHO

OHH

HOCH2

H

O

TRANSFERASESGlycosyltransferase – if the transferred

group is a carbohydrate residue

Page 51: Enzymes and enzyme kinetics 2012

HYDROLASESCatalyze cleavage of chemical bonds by

addition of H2O, producing 2 products

O O || ||-O – P ~ O – P ~ O- + HOH | | -O O-

Pyrophosphate (PPi)

O || 2 HO – P – O- | -O Phosphate 2 (Pi)

Pyrophosphatase

Phosphate bond

Page 52: Enzymes and enzyme kinetics 2012

O O || ||CH3-(CH2)12-C-CH2-C-S-CoA

O || CH3-(CH2)12-C-S-CoA

β-Ketoacyl CoA

Fatty acyl CoA (2 carbons shorter)

Thiolase CH3-C-S-CoA

CoA

Acetyl CoA

HYDROLASESCatalyze cleavage of chemical bonds,

producing 2 products

Page 53: Enzymes and enzyme kinetics 2012

LYASES

Cleave C-C, C-O, C-N bonds by means other than hydrolysis or oxidation

O O-

\\ / C | C = O H+

| CH3

Pyruvate

H O \ // C + O = C = O | Carbon CH3 dioxide (CO2)

Pyruvatedecarboxylase

Acetaldehyde

Page 54: Enzymes and enzyme kinetics 2012

LYASES

Dopamine

3,4-Dihydroxyphenylalanine (DOPA)

CH2 – CH – COO-

I NH3

+

OH

HO

CH2 – CH2 – NH3

OH

HO

+

DOPA decarboxylaseCO2

43

PLP

Cleave C-C, C-O, C-N bonds by means other

than hydrolysis or oxidation

Page 55: Enzymes and enzyme kinetics 2012

H|

CH2 – C – COO-

| NH3

PLP

CH2 – CH2 – NH3

Histidinedecarboxylase

Histidine

HISTAMINE

CO2

LYASESCleave C-C, C-O, C-N bonds by means other

than hydrolysis or oxidation

Page 56: Enzymes and enzyme kinetics 2012

Catalyze C-C bond cleavage in a reversible reaction

GLYCOLYSIS

P P | | O O H OH OH O | || | | | |H - C1 – C2 – C3 - C4 - C5 - C6 - H | | | | | H OH H H HFructose 1,6-Bisphosphate

O ||H – C1 – H |H – C2 – OH |H – C3 – O – P | H

O ||H – C4 – O – P |H – C5 – OH |H – C6 – OH | H

Glyceraldehyde3 – Phosphate (GADP)

DihydroacetonePhosphate (DHAP)

Aldolase A

Δ G0 = + 5.73 kcal/mole

LYASES

Page 57: Enzymes and enzyme kinetics 2012

LYASESSynthase – catalyzes a physiologically important reaction that favors the formation of a C-C bond

S~CoA|C1 = O|C2H3

C1OO-

|C2= O|C3H2

|C4OO-

C1OO-

| C2H2

| HO – C3 – C4OO-

| C5H2

| C6OO-

Oxaloacetate (C4)

Acetyl CoA (C2) Citrate (C6)

H2O HS-CoA

Citrate synthase

Page 58: Enzymes and enzyme kinetics 2012

LYASESSynthase – catalyzes a physiologically important reaction that favors the formation of a C-C bond

HS-CH2-CH2-CH-COO-

I NH3

+

CH2OHI

HC-NH3+

I COO-

Homocysteine

Serine CH2

I CH2

I CH-NH3

+

I COO-

CH2

I H -C-NH3

+

I COO-

S

Cystathione

Cystathionine synthase

H2O

PLP

Page 59: Enzymes and enzyme kinetics 2012

LYASES

Hydratase – add H2O to a susbtrate

C1OO-

| H – C2

|| C3– H | C4OO-

Fumarate

C1OO-

|HO – C2 – H | H – C3 – H | C4OO-

Malate

Fumarase(or fumarate hydratase

H2O

Page 60: Enzymes and enzyme kinetics 2012

ISOMERASESTransfer of functional groups or double

bonds within the same molecule

C1OO-

|H3N – C2 – H

| C3H3

L-Alanine

C1OO-

| H – C2 – NH3

| C3H3

D-Alanine

Alanineracemase

Page 61: Enzymes and enzyme kinetics 2012

Transfer of functional groups or double bonds within the same molecule

GLYCOLYSIS

O ||H – C1 – H |H – C2 – OH |

H – C3 – O – P | H

O ||

H – C1 – O – P |H – C2 – OH |H – C3 – OH | H

Glyceraldehyde3 – Phosphate (GADP)

DihydroacetonePhosphate (DHAP)

ISOMERASES

Triosephosphateisomerase

Page 62: Enzymes and enzyme kinetics 2012

(Aldose)(Ketose)

O ||

C1 - H

| H - C2 - OH |OH - C3 - H | H - C4 - OH | H - C5 - OH | H - C6 - O – P | H

H | H – C1 - OH |

C2 = O

|OH - C3 - H | H - C4 - OH | H - C5 - OH | H - C6 - O – P | H

Phosphohexoisomerase

Aldehyde group

Keto group

Glucose 6-Phosphate Fructose 6-Phosphate

ATP

ADP

ISOMERASESTransfer of functional groups or double

bonds within the same molecule

Page 63: Enzymes and enzyme kinetics 2012

O || C1 – O-

|H – C2 – O - P |H – C3 – OH | H

O || C1 – O-

|H – C2 – OH |H – C3 – O – P | H

3-Phosphoglycerate 2-Phosphoglycerate

Phosphoglycerate mutase

Mg+2

Δ G0 = + 1.06 kcal/mole

ISOMERASESTransfer of functional groups or double

bonds within the same molecule

Page 64: Enzymes and enzyme kinetics 2012

ISOMERASESTransfer of functional groups or double

bonds within the same molecule

C1H2OH I

C2=O I

HOC3H I

HC4OH I

H2C5OPO32-

C1H2OH I

C2=O I

HC3OH I

HC4OH I

H2C5OPO32-

Epimerase

D- Xylulose 5-phosphate D- Ribulose 5-phosphate

C-3 Epimers

Page 65: Enzymes and enzyme kinetics 2012

CH3

|C = O |COO-

COO-

|CH2

|C = O |COO-Biotin-

CO2

ATPADP + Pi

OxaloacetatePyruvate

Pyruvate carboxylase

ATP ADP+ Pi

LIGASESCatalyze the joining of

substrates in the presence of ATP.

Page 66: Enzymes and enzyme kinetics 2012

CHARACTERISTICS OF ENZYMES

They are not changed by the reaction they catalyze.

Page 67: Enzymes and enzyme kinetics 2012

They do not change or alter the equilibrium of the

chemical reaction.

CHARACTERISTICS OF ENZYMES

Page 68: Enzymes and enzyme kinetics 2012

They increase reaction rates by decreasing

the activation

energy.

CHARACTERISTICS OF ENZYMES

Page 69: Enzymes and enzyme kinetics 2012

ENZYMES DECREASE THEACTIVATION ENERGY

Reaction progress

ΔGfor the reaction

ΔG

+

++(catalyzed)

ΔG++

(uncatalyzed)

Transition state, S

++

Substratesor

Reactants(e.g. CO2 + H2O)

Products(H2CO3)F

ree

en

erg

y, ∆

GReaction Coordinate Diagram

Page 70: Enzymes and enzyme kinetics 2012

They are highly specific for the reactants or substrates

they act on and catalyze only one type

of chemical

reaction.

CHARACTERISTICS OF ENZYMES

Page 71: Enzymes and enzyme kinetics 2012

ENZYME SPECIFICITY

CO2 + H2OH2CO3

Carbonicanhydrase

Page 72: Enzymes and enzyme kinetics 2012

ENZYME SPECIFICITY

Catalase

2 H2O2 2 H2O

Page 73: Enzymes and enzyme kinetics 2012

ENZYME SPECIFICITY COO-

| HO - C – H | CH3

COO-

I C – OH

I CH3

Lactate dehydrogenase

L-Lactate PyruvateNAD+ NADH+ + H+

COO-

| H - C – OH | CH3

D-Lactate

Page 74: Enzymes and enzyme kinetics 2012

ENZYME SPECIFICITY O || C1 - H | H - C2 - OH |OH - C3 - H | H - C4 - OH | H - C5 - OH |

H - C6 – OH

| H

Glucose

O || C1 - H | H - C2 - OH |OH - C3 - H | H - C4 - OH | H - C5 - OH |

H - C6 - O – P

| HGlucose 6-phosphate

Glucokinase

ATP ADP + Pi

Mg+2

Page 75: Enzymes and enzyme kinetics 2012

CHARACTERISTICS OF ENZYMES

HI

R – C – CO –I

NH2

H I

NH – C – R’ I

COOH

PEPTIDE BOND

They are mostly proteins in nature.

Page 76: Enzymes and enzyme kinetics 2012

ENZYME-SUBSTRATE COMPLEX FORMATIONThe First Step in Enzymatic Catalysis

Substrate concentration [S]

Rea

ctio

n v

elo

city

(V

)

Maximal velocity

Page 77: Enzymes and enzyme kinetics 2012

ENZYME-SUBSTRATE COMPLEXCYTOCHROME P450-CAMPHOR

Camphor

Cytochrome P-450

Page 78: Enzymes and enzyme kinetics 2012

MODELS OF ENZYME-SUBSTRATE COMPLEX

Lock and Key Model (Emil Fischer, 1894) Induced Fit Model (Daniel E. Koshland, Jr, 1958)

Page 79: Enzymes and enzyme kinetics 2012

LOCK AND KEY MODEL

Page 80: Enzymes and enzyme kinetics 2012

INDUCED FIT MODEL

Page 81: Enzymes and enzyme kinetics 2012

KINETICS OF ENZYME-CATALYZED REACTIONS

E + S ES E + Pk1

k-1

k2

k-2

Substrate binding

Catalyticstep

Page 82: Enzymes and enzyme kinetics 2012

MICHAELIS-MENTEN EQUATION

Vo = Vmax [S]{Km + [S]}

Vo = Velocity at any time (moles/time)Vmax = Maximal velocity (or reaction rate)Km = Michaelis constant for the particular enzyme under investigation = (K-1 + K2)/K1

[S] = Substrate concentration (molar)

Page 83: Enzymes and enzyme kinetics 2012

MICHAELIS-MENTEN SATURATION CURVE

Substrate concentration [S]

Km

Vmax

2

Vmax

Rea

ctio

n v

elo

city

(V

O)

Zero order

B

A

C

First order

│││││││ ││

││

││

││

A = [S] < Km

B = [S] = Vmax/2

C = [S] > Km

= Vo is maximal(Vmax)

Page 84: Enzymes and enzyme kinetics 2012

A B C

=S

=E

REPRESENTATION OF AN ENZYME IN THE PRESENCE OF A SUBSTRATE

[S] < Km [S] = Vmax/2 [S] > Km

Page 85: Enzymes and enzyme kinetics 2012

1. It is the substrate concentration at which half of the active sites of the enzyme are filled up.

2. It is an inverse measure of the affinity of the substrate for the enzyme:

a. The lower the Km, the higher is the

affinity.

b. The higher the Km, the lower is the affinity.

SIGNIFICANCE OF KM

Page 86: Enzymes and enzyme kinetics 2012

VI. Reactions: GLYCOLYSIS

ATP ADP + Pi

Mg+2

O || C1 - H | H - C2 - OH |OH - C3 - H | H - C4 - OH | H - C5 - OH | H - C6 – OH | H

Glucose

O || C1 - H | H - C2 - OH |OH - C3 - H | H - C4 - OH | H - C5 - OH | H - C6 - O – P | HGlucose 6-

phosphate

KM AND PHYSIOLOGICAL UTILIZATION OF GLUCOSE

Hexokinase(Extrahepatic cells, RBCs)

Glucokinase(Liver, Pancreatic β-cells

Km for hexokinase = 0.1 mMKm for glucokinase = 5 mM

Page 87: Enzymes and enzyme kinetics 2012

LINEWEAVER- BURKE DOUBLE RECIPROCAL PLOT

1

[S]

1V

Intercept on Y-axis = 1

Vmax

Intercept on X-axis =

Slope = Km

Vmax

Km

- 1

Page 88: Enzymes and enzyme kinetics 2012

LINEWEAVER-BURKE DOUBLE RECIPROCAL PLOT:SAMPLE PROLEM

I I III I II II I I I.050 .25.1 .1

5.2

.3

55

50

45

35

40

30

25

20

15

10

1/[S] (mM-1)

1/V

(m

M s

ec

-1)-

1

Y intercept = 1

Vmax

X intercept = 1

Km

Slope = KmVmax

-

- .12

Y intercept

I IIII

Page 89: Enzymes and enzyme kinetics 2012

LINEWEAVER-BURKE DOUBLE RECIPROCAL PLOT: SAMPLE PROBLEM

Y intercept = 1

Vmax

Vmax =1

Y intercept

Vmax =1

20

Vmax = 0.05 (mM sec-1)-1

X intercept = -1

Km = -

--

Km

1

.12Km =

8.33 (mM-1)

X intercept

1

Km =

Page 90: Enzymes and enzyme kinetics 2012

INHIBITION OF ENZYMATIC REACTIONS

Reversible

a. Competitive

b. Non-competitive

c. Uncompetitive

Irreversible

Page 91: Enzymes and enzyme kinetics 2012

REVERSIBLE INHIBITION

Page 92: Enzymes and enzyme kinetics 2012

COMPETITIVE INHIBITION: LOVASTATIN

Page 93: Enzymes and enzyme kinetics 2012

Inhibitor Type Binding Site on Enzyme Kinetic Effect

Competitive Inhibitor

Inhibitor binds specifically at active or catalytic site, where it competes with substrate for binding; inhibition is reversed by increasing substrate concentration.

Vmax unchanged; Km increased to reach a given velocity.

Noncompetitive Inhibitor

Inhibitor binds E or ES other than at the active or catalytic site; substrate binding unaltered but ESI complex cannot form due to structural change in the enzyme →↓ catalytic power no products formed; inhibition cannot be reversed by increasing substrate concentration since inhibitor cannot be driven from the enzyme.

Vmax decreased proportionately to inhibitor concentration; Km unchanged (since substrate can still bind to the enzyme).

Uncompetitive Inhibitor

Inhibitor binds only to ES complexes at locations other than catalytic site; substrate binding modifies enzyme structure, making inhibitor-binding site available; inhibition cannot be reversed by increasing substrate concentration; rare in occurrence.

Vmax decreased

Km decreased

REVERSIBLE INHIBITON

Page 94: Enzymes and enzyme kinetics 2012

Vmax unchanged by competitive inhibitors since increasing substrate concentration can displace virtually all competitive inhibitors bound to theactive site, hence the identical Y-axis intercepts of Lineweaver-Burke plots, with or without inhibitor.

A competitive inhibitor increases Km for a given substrate in order to attain Vmax that were reached in its absence, hence the differing –X intercepts.

1[S]

1[ V ]

Uninhibited enzyme

Competitiveinhibitor

1Vmax

LINEWEAVER-BURKE PLOT FOR COMPETITIVE INHIBITION

-1Km

Page 95: Enzymes and enzyme kinetics 2012

Vmax is decreased since a

noncompetitive inhibitor

reduces the concentration of

ES complex that can advance

to reaction products, hence

the differing Y-intercepts.

Km is unchanged since

substrate can still bind to the

enzyme, hence the identical

–X intercepts of Lineweaver-

Burke plots, with or without

noncompetitive inhibitor. 1

[S]

1[ V ]

Uninhibited enzyme

Noncompetitiveinhibitor

LINEWEAVER-BURKE PLOT FOR NONCOMPETITIVE INHIBITION

- 1 Km

1Vmax

Page 96: Enzymes and enzyme kinetics 2012

Uncompetitive inhibitors

decrease both Vmax and

Km, hence the

production of parallel

lines in uninhibited and

inhibited reactions.

1[S]

1[ V ]

Uninhibited enzyme

Uncompetitiveinhibitor

LINEWEAVER-BURKE PLOT FOR UNCOMPETITIVE INHIBITION

1 Km

-

1Vmax

Page 97: Enzymes and enzyme kinetics 2012

LINEWEAVER-BURKE DOUBLE RECIROCAL PLOT IN THE PRESENCE OF AN INHIBITOR

1[S]

1[ V ]

1Vmax

Noncompetitive inhibitor

Competitive inhibitor

Uninhibitedenzyme

Uncompetitive inhibitor

-1Km

Type of Inhibition

Vmax

Km

CompetitiveSame ↑

Noncompetitive↓ Same

Uncompetitive↓ ↓

Page 98: Enzymes and enzyme kinetics 2012

IRREVERSIBLE INHIBITION Diisopropylphosphofluoridate (DIPF)

OII

H3C-C- O-CH2-CH2-N(CH3)3

OH I

Enz-Ser

+ HO- CH2-CH2-N(CH3)3

O I

Enz-Ser

OII

- C – CH3

Acetylcholine Choline

+

OH I

Enz-Ser

H2O

O II

H3C-C-O-

Acetate

A. Normal Reaction of Acetylcholinesterase

A. Reaction with Organophosphorus Inhibitors

OH I

Enz-Ser +

CH3

I H-C-O -

I CH3

CH3

IO-C-O-

I CH3

O II

P - I

F F-, H+

CH3

I H-C-O -

I CH3

CH3

IO-C-O-

I CH3

O II

P - I

O

Enz-Ser

Inactiveenzyme

+

(Enz – acetylcholinesterase)

Page 99: Enzymes and enzyme kinetics 2012

IRREVERSIBLE INHIBITION - Penicillin

COO-

CH3

N

CS

C

CCH3

H

Penicillin I C=O I H-N I H-C – I C II O

H

I C=O I H-N I H-C – I O=C II

COO-

CH3

N

CS

C

CCH3

H

H

OHI

Ser

Glycopeptidetranspeptidase

OI

Ser

Glycopeptidetranspeptidase

H

Bacterial enzyme

Β-lactam ring

Strained peptide bond

Page 100: Enzymes and enzyme kinetics 2012

REPRESENTATIVE DRUGS THAT INHIBIT SPECIFIC ENZYMES

DRUG ENZYME TARGET DISEASE

Amrubicin® Topoisomerase II CA chemotherapy

Antabuse® Aldehyde dehydrogenase

Alcoholism

Captopril® Angiotensin-converting enzyme

Hypertension

Celebrex® Cyclooxyenase-2 Arthritis

Digoxin® Na+-K+-ATPase pump Heart problem

Agenerase® HIV protease Acquired Immunodeficiency Syndrome (AIDS)

Lipitor® HMG CoA reductase Hypercholesterolemia

Viagra® Phosphodiesterase Erectile dysfunction

Page 101: Enzymes and enzyme kinetics 2012

ENZYMES AND PHARMACOTHERAPY: ACE INHIBITORS

Angiotensinogen(liver, polypeptide,

400 AAs)

Renin(JG cells, placenta)

Angiotensin I(decapeptide, 10 AAs)

Angiotensin II(octapeptide, 8 AAs)

Angiotensin converting

enzyme (ACE)

Functions/Effects

Arteriolar vasocons-

triction

Aldosterone-mediated renal

Na+ & H2O reabsorption

↑ BP

ACE Inhibitor (Captopril, Amlodipine)

_

Page 102: Enzymes and enzyme kinetics 2012

HMG CoA (3-hydroxy-3-methylglutaryl CoA )

2 NADPH + 2H+

2 NADP+

HMG CoA reductase

O OH II | -O – C – CH2 – C – CH2 – CH2OH | CH3

MEVALONATE (C6)

Statins Atorvastatin(Ex. Lipitor)

CoA

O OH O || | || C – CH2 – C – CH2 – C – S-CoA / |O- CH3

ENZYMES AND PHARMACOTHERAPY: STATINS

-

CHOLESTEROL

Page 103: Enzymes and enzyme kinetics 2012

Viagra® (Sildenafil citrate)

GMP

Smooth muscle relaxation

and vasodilation in penile

blood vessels

SUSTAINED ERECTION

GTP cGMP

Adenylate cyclase

Phospho-diesterase

ENZYMES AND PHARMACOTHERAPY: VIAGRA®

Pi

↑ cGMP

-

Page 104: Enzymes and enzyme kinetics 2012

ENZYMES AND PHARMACOTHERAPY:COX-2 INHIBITORS

PHOSPHOLIPIDS (Cell Membrane)

Arachidonic acid (C20:Δ4) (Eicosatetraenoic Acid)

5-Lipoxygenase Cyclooxy-genase

PGI2

(Prostacyclin)

TXA2

(ThromboxaneA2)

PGE2

PGF2α

Phospholipase A2

LTC4 LTD4

LTB4

LTE4

LTA4

(Leukotriene A4)

Glu

Glutathione(Glu-Gly-Cys)

ProstaglandinH2 synthase

ProstaglandinH2 synthase

PGG2

(Prostaglandin G2)

Peroxidase

PGH2

(Prostaglandin H2)Prostacyclin

synthaseThromboxane

synthaseIsomerase

Reductase

5-HPETE 5-HETESpontaneousPeroxidase

Leukotriene synthase

Leukotriene synthase

Gly

COX-1 COX-2

-NSAIDS-Selective COX-2 inhibitors (Ex. Celebrex®)

NSAIDS (ASA, Indomethacin)

Pain and inflammation

EICOSANOID SYNTHESIS

--

Page 105: Enzymes and enzyme kinetics 2012

ENZYMES AND PHARMACOTHERAPY: ANTABUSE®

CH2 – CH2 – OHEthanol

CH3 - OHAcetaldehyde

Alcohol dehydrogenaseNAD+

NADH + H+

Aldehyde dehydrogenase

Acetate

Acetyl CoA

Antabuse®

_

Page 106: Enzymes and enzyme kinetics 2012

KINETICS FOR AN ALLOSTERIC ENZYME

Page 107: Enzymes and enzyme kinetics 2012

REGULATION OF ENZYME ACTIVITY

Feedback Inhibition Allosteric (Non-covalent) Modification Covalent Modification Zymogen Activation Induction or Repression of Enzyme Synthesis

Page 108: Enzymes and enzyme kinetics 2012

FEEDBACK INHIBITIONOriginal Precursor(s)

Enzyme 1

Enzyme 2

Enzyme 3

Enzyme 4

Enzyme 5

1

2

3

4

Final Products

Page 109: Enzymes and enzyme kinetics 2012

FEEDBACK INHIBITION

Carbamoyl PO4 + Aspartate

Carbamoyl aspartate

Aspartate transcarbamoylase (ATCase)

series of reactions

Cytidine triphosphate (CTP)

RNA & DNA synthesis

Page 110: Enzymes and enzyme kinetics 2012

FEEDBACK INHIBITION OF HMG CoA REDUCTASE BY CHOLESTEROL

Acetyl CoA

Acetyl CoA

HMG CoA

Mevalonic acid + CoA

HMG CoA reductase

Cholesterol

Feedbackinhibition

Acetoacetyl CoA

2NADPH

+ 2H

+

2NA

DP

++

2H+

> 25 steps

Page 111: Enzymes and enzyme kinetics 2012

ALLOSTERIC MODIFICATIONAllosteric modulator (activator or inhibitor)

Binds to regulatory or allosteric site

Conformational change in the regulatory enzyme

Effect is transmitted to the active site

Change in shape of the active site

Altered activity

Page 112: Enzymes and enzyme kinetics 2012

ALLOSTERIC MODIFICATION:Phosphofructokinase I

Page 113: Enzymes and enzyme kinetics 2012

COVALENT MODIFICATION

ATP ADP

ENZYME- Ser -- OH

HPO4= H2O

Proteinkinase

Phospho-protein

phosphatase

ENZYME- Ser – O – PO32-

Page 114: Enzymes and enzyme kinetics 2012

COVALENT MODIFICATION OF THE ENZYME

Glycogen phosphorylase

AMPATP

and/orG6P

Glucose

2 ATP 2 ADP

2 H2O2 P

Phosphorylasekinase

Phosphorylase b(inactive)

Phosphorylase a(active)

PP

P P

Phosphoproteinphosphatase

Page 115: Enzymes and enzyme kinetics 2012

COVALENT MODIFICATION: PYRUVATE DEHYDROGENASE

Pyruvate dehydrogenase

Pyruvatedehydrogenase

P

Pyruvate dehydrogenase

kinase

ATP ADP

Pyruvate dehydrogenase

phosphatasePi H2O

(inactive)(active)

Page 116: Enzymes and enzyme kinetics 2012

ENZYMES Low activity High activity

Acetyl CoA Carboxylase EP E

Glycogen synthase EP E

Pyruvate dehydrogenase EP E

HMG CoA reductase EP E

Glycogen phosphorylase E EP

Citrate lyase E EP

Phosphorylase b kinase E EP

HMG CoA reductase kinase E EP

MAMMALIAN ENZYMES WHOSE CATALYTIC ACTIVITY IS ALTERED BY COVALENT PHOSPHORYLATION-DEPHOSPHORYLATION

E = Dephosphorylated EP = Phosphoenzyme

Page 117: Enzymes and enzyme kinetics 2012

ZYMOGEN ACTIVATIONChymotrypsinogen

Page 118: Enzymes and enzyme kinetics 2012

ZYMOGEN ACTIVATION: BLOOD COAGULATION

Clotting Factors

Prothrombin Thrombin

Fibrinogen Fibrin

Ca+2

Some of the processes involved in blood clotting

Page 119: Enzymes and enzyme kinetics 2012

INDUCTION OR REPRESSIONOF ENZYME SYNTHESIS

↑ Blood glucoselevels

(Well-fed state)

↑ Insulin

↑ Synthesis of key enzymes involvedin glucose degradation

↓ Blood glucose levels(Starvation)

↑ Glucagon

↑ Synthesis of key enzymes involvedin glucose synthesis

Page 120: Enzymes and enzyme kinetics 2012

FACTORS AFFECTING ENZYME ACTIVITY

Temperature pH Substrate concentration Co-factors

Page 121: Enzymes and enzyme kinetics 2012

EFFECT OF TEMPERATURER

eacti

on

velo

cit

y (V

o)

Temperature (oC)

││ ││ ││ ││ │ ││ │ ││ ││

││

││

││

││

││

││

70605040302010 80

Optimum T

Heat inactivation

of the enzyme

Increasing enzyme activity

Page 122: Enzymes and enzyme kinetics 2012

EFFECT OF pH

Optimum pH

Page 123: Enzymes and enzyme kinetics 2012

OPTIMUM pH OF VARIOUS ENZYMES

Page 124: Enzymes and enzyme kinetics 2012

EFFECT OF CO-FACTORS: Chlorides, Bromides, Iodides

Cofactors increase the rate of enzyme-catalyzed reactions

Page 125: Enzymes and enzyme kinetics 2012

EFFECT OF SUBSTRATE CONCENTRATION

Substrate concentration [S]

Vmax

Re

act

ion

ve

loc

ity

(V

)

Page 126: Enzymes and enzyme kinetics 2012

ENZYMES IN CLINICAL DIAGNOSIS

Page 127: Enzymes and enzyme kinetics 2012

CARDIAC ENZYMES AS MARKERS FORMYOCARDIAL INFARCTION (MI)

Aspartate aminotransferase

Page 128: Enzymes and enzyme kinetics 2012

QUESTION 1

Which of the following is TRUE when a substrate concentration equals km in an enzyme-catalyzedreaction? A. A few of the enzyme molecules are present as ES complex.B. Majority of the enzyme molecules are present as ES complex.C. Half of the enzyme molecules are present as ES complex.D. All of the enzyme molecules are present as ES complex.

Page 129: Enzymes and enzyme kinetics 2012

QUESTION 1

Which of the following is TRUE when a substrate concentration equals km in an enzyme-catalyzedreaction? A. A few of the enzyme molecules are present as ES complex.B. Majority of the enzyme molecules are present as ES complex.C. Half of the enzyme molecules are present as ES complex.D. All of the enzyme molecules are present as ES complex.

Page 130: Enzymes and enzyme kinetics 2012

Competitive inhibition can be relieved by

increasing which of the following?

A. Enzyme concentration

B. Inhibitor concentration

C. Enzyme-substrate concentration

D. Substrate concentration

QUESTION 2

Page 131: Enzymes and enzyme kinetics 2012

Competitive inhibition can be relieved by

increasing which of the following?

A. Enzyme concentration

B. Inhibitor concentration

C. Enzyme-substrate concentration

D. Substrate concentration

QUESTION 2

Page 132: Enzymes and enzyme kinetics 2012

Which of the following enzymes requires

biotin as a coenzyme?

A. PEP carboxykinase

B. Pyruvate carboxylase

C. Phosphofructokinase I

D. Pyruvate dehydrogenase

QUESTION 3

Page 133: Enzymes and enzyme kinetics 2012

Which of the following enzymes requires

biotin as a coenzyme?

A. PEP carboxykinase

B. Pyruvate carboxylase

C. Phosphofructokinase I

D. Pyruvate dehydrogenase

QUESTION 3

Page 134: Enzymes and enzyme kinetics 2012

An enzyme with a low Km indicates which of the following? A. High affinity for the substrate.B. Requires increased amount of substrate to become saturated.C. Vmax can be reached at high substrate concentration.D. Less enzyme-substrate complexes are formed.

QUESTION 4

Page 135: Enzymes and enzyme kinetics 2012

An enzyme with a low Km indicates which of the following? A. High affinity for the substrate.B. Requires increased amount of substrate to become saturated.C. Vmax can be reached at high substrate concentration.D. Less enzyme-substrate complexes are formed.

QUESTION 4

Page 136: Enzymes and enzyme kinetics 2012

QUESTION 5

Enzymes can be regulated when

phosphorylated. This type of regulation

is called:

A. Allosteric control

B. Feed back regulation

C. Covalent modification

D. Enzyme induction

Page 137: Enzymes and enzyme kinetics 2012

QUESTION 5

Enzymes can be regulated when

phosphorylated. This type of regulation

is called:

A. Allosteric control

B. Feed back regulation

C. Covalent modification

D. Enzyme induction

Page 138: Enzymes and enzyme kinetics 2012

REFERENCES

• Lehninger’s Principles of Biochemistry, Nelson, D.L., Cox, M.M., 5th ed., pp. 183-220. • Harper’s Illustrated Biochemistry, Murray, R. K., et. al., 29th ed., pp. 62-82. • Biochemistry with Clinical Correlations, Devlin, M. T., 7th ed., pp. 378-421.• Biochemistry, Lippincott’s Illustrated Reviews, Champe, P.C., Harvey, R.A., 4th ed., pp. 53- 67. • Principles of Biochemistry, Horton, H.R., et al., 4th ed., pp. 129-156. • Marks’ Basic Medical Biochemistry: A Clinical Approach, Lieberman, M., Marks, A.D., 3rd ed., pp. 116-154.

Page 139: Enzymes and enzyme kinetics 2012

THANK YOUTHANK YOU