Gastro-intestinal EnzymesGastro-intestinal Enzymes

PepsinPepsin

• Proteolytic enzyme of the stomachProteolytic enzyme of the stomach• At least two immunologically distinct pepsinogens (1 and At least two immunologically distinct pepsinogens (1 and

2) are secreted by the stomach2) are secreted by the stomach• Pepsinogen 1 is secreted by the chief cells in the oxyntic Pepsinogen 1 is secreted by the chief cells in the oxyntic

glandular areaglandular area• Pepsinogen 2 by cells through out the stomach as well as in Pepsinogen 2 by cells through out the stomach as well as in

Brunner’s glands in the duodenumBrunner’s glands in the duodenum• It is an endopeptidaseIt is an endopeptidase• It preferentially hydrolyses peptide linkage where one of It preferentially hydrolyses peptide linkage where one of

the amino acids is aromatic (eg, tyrosin) or dicarboxylic the amino acids is aromatic (eg, tyrosin) or dicarboxylic amino acids (eg, glutamate)amino acids (eg, glutamate)

• Pepsin potentiates rather than initiates ulcer formationPepsin potentiates rather than initiates ulcer formation

Clinical SignificanceClinical Significance

• Serum concentration of pepsinogen I reflects the parietal Serum concentration of pepsinogen I reflects the parietal cell mass and correlate well with the maximum acid-cell mass and correlate well with the maximum acid-secreting capacitysecreting capacity

• Increased pepsinogen I:Increased pepsinogen I:– Duodenal ulcer Duodenal ulcer – Acute and chronic superficial gastritis. Acute and chronic superficial gastritis. – H. pylori sero-positive patientsH. pylori sero-positive patients

• Decreased levels of pepsinogen I:Decreased levels of pepsinogen I:– Atrophic gastritis Atrophic gastritis – Gastric carcinomaGastric carcinoma

• Increased pepsinogen II:Increased pepsinogen II:– Acute and chronic superficial gastritisAcute and chronic superficial gastritis

• Major risk factor for gastric ulcerMajor risk factor for gastric ulcer

Pancreatic JuicePancreatic Juice

• Alkaline secretion:Alkaline secretion: secreted by the duct cellssecreted by the duct cells– Composition: HCOComposition: HCO33

--, Na, Na++, K, K++, Mg, Mg22++, Ca, Ca22

++, Cl, Cl--

– Function: together with the other alkaline secretions (bile and Function: together with the other alkaline secretions (bile and intestinal juice) neutralize the acid chyme arriving from the intestinal juice) neutralize the acid chyme arriving from the stomach. stomach.

Why this is important?Why this is important?• The pancreatic enzymes require a neutral or slightly alkaline The pancreatic enzymes require a neutral or slightly alkaline

pH for their activitypH for their activity• The absorption of fat depends on the formation of micelles, a The absorption of fat depends on the formation of micelles, a

process which only takes place at neutral or slightly alkaline process which only takes place at neutral or slightly alkaline pHpH

• It protects the intestinal mucosa, excess acid in the duodenum It protects the intestinal mucosa, excess acid in the duodenum can damage the mucosa and can lead to ulcer formationcan damage the mucosa and can lead to ulcer formation

Pancreatic JuicePancreatic Juice

• Pancreatic enzymes: secreted by acinar cellsPancreatic enzymes: secreted by acinar cells– In their inactive form;In their inactive form;

• TrypsinogenTrypsinogen• ChymotrypsinogenChymotrypsinogen• ProelastaseProelastase• ProcarboxypeptidaseProcarboxypeptidase

• Prophospholipase AProphospholipase A22

– In their active form;In their active form;• LipaseLipase• Alpha-amylaseAlpha-amylase• RibonucleaseRibonuclease• DeoxyribonucleaseDeoxyribonuclease• Cholesterol esteraseCholesterol esterase

TrypsinTrypsin

• Two types, Two types, trypsinogen-1 and -2trypsinogen-1 and -2• Trypsin is a serine proteinase charaterized by the presence at Trypsin is a serine proteinase charaterized by the presence at

the active site of serine and histidine, both of which participate the active site of serine and histidine, both of which participate in the catalytic processin the catalytic process

• Trypsins are considered endopeptidasesTrypsins are considered endopeptidases • The enzymatic mechanism is like all other serine proteases: A The enzymatic mechanism is like all other serine proteases: A

catalytic triadcatalytic triad serves to make the serves to make the active siteactive site serineserine nucleophilic. nucleophilic. This is achieved by modifying the electrostatic environment of This is achieved by modifying the electrostatic environment of the serine residuethe serine residue

• The The aspartateaspartate residue (Asp 189) located in the catalytic pocket residue (Asp 189) located in the catalytic pocket (S1) of trypsins is responsible for attracting and stabilizing (S1) of trypsins is responsible for attracting and stabilizing positively-charged positively-charged lysinelysine and/or and/or argininearginine, and is thus responsible , and is thus responsible for the specificity of the enzyme.for the specificity of the enzyme.

TrypsinTrypsin

• Trypsin activity is stimulated by calcium and magnesium Trypsin activity is stimulated by calcium and magnesium ions and to a lesser extent by cobalt and manganese ions . ions and to a lesser extent by cobalt and manganese ions . Cyanide, sulfide, citrate, fluoride, and heavy metals inhibit Cyanide, sulfide, citrate, fluoride, and heavy metals inhibit activity as do those organic phosphorous compounds that activity as do those organic phosphorous compounds that combine with serine at the active site. combine with serine at the active site.

• Clinical Significance:Clinical Significance: – deficiency of trypsin might lead to the disorder termed deficiency of trypsin might lead to the disorder termed

meconium ileusmeconium ileus– After an attack of acute pancreatitis, serum After an attack of acute pancreatitis, serum

immunoreative trypsin rises in parallel with serum immunoreative trypsin rises in parallel with serum amylase activity.amylase activity.

ChymotrypsinChymotrypsin

• Two types, Two types, chymotrypsinogenchymotrypsinogen-1 and -2-1 and -2

• Chymotrypsin is synthesized in the acinar Chymotrypsin is synthesized in the acinar cells of the human cells of the human pancreaspancreas

• Chymotrypsin-1 describes as anionic and Chymotrypsin-1 describes as anionic and chymotrypsin-2 as cationic because of their chymotrypsin-2 as cationic because of their differing electrophoretic mobilities; the differing electrophoretic mobilities; the cationic form predominates.cationic form predominates.

Action and kinetics of chymotrypsin:Action and kinetics of chymotrypsin:

• For an enzyme-mediated reaction to take place, the For an enzyme-mediated reaction to take place, the reacting molecule or molecules, called substrates, must fit reacting molecule or molecules, called substrates, must fit into a specific section of the enzymes structure called the into a specific section of the enzymes structure called the active site.active site.

• Each active site has:Each active site has:(1) a shape that fits a specific substrate or substrates(1) a shape that fits a specific substrate or substrates(2) side chains that attract the enzymes particular (2) side chains that attract the enzymes particular substratessubstrates(3) side chains specifically positioned to speed the reaction.(3) side chains specifically positioned to speed the reaction.

Mechanism of peptide bond cleavage in chymotrypsin (1)Mechanism of peptide bond cleavage in chymotrypsin (1)

Mechanism of peptide bond cleavage in chymotrypsin (2)Mechanism of peptide bond cleavage in chymotrypsin (2)

Mechanism of peptide bond cleavage in chymotrypsin (3)Mechanism of peptide bond cleavage in chymotrypsin (3)

Mechanism of peptide bond cleavage in chymotrypsin (4)Mechanism of peptide bond cleavage in chymotrypsin (4)

Mechanism of peptide bond cleavage in chymotrypsin (5)Mechanism of peptide bond cleavage in chymotrypsin (5)

Mechanism of peptide bond cleavage in chymotrypsin (6)Mechanism of peptide bond cleavage in chymotrypsin (6)

ChymotrypsinChymotrypsin

• Clinical Significance:Clinical Significance: – Chymotrypsin is more resistant than Chymotrypsin is more resistant than

trypsin to degradation in the intestine; it trypsin to degradation in the intestine; it is therefore the enzyme of choice for is therefore the enzyme of choice for assay in feces.assay in feces.

– The major application of chymotrypsin The major application of chymotrypsin assay is the investigation of pancreatic assay is the investigation of pancreatic disease.disease.

Acute PancreatitisAcute Pancreatitis

• Acute pancreatitis is a disease in which the pancreatic Acute pancreatitis is a disease in which the pancreatic tissue is destroyed by digestive enzymestissue is destroyed by digestive enzymes

• The pancreas normally secrets a polypeptide known as The pancreas normally secrets a polypeptide known as Kazal inhibitor, that inhibits any small amounts of Kazal inhibitor, that inhibits any small amounts of activated trypsin which may find its way into the ducts, by activated trypsin which may find its way into the ducts, by forming a complex with itforming a complex with it

• Enzyme Y is also exhibiting a protective function, which is Enzyme Y is also exhibiting a protective function, which is activated by traces of active trypsin degrades zymogenactivated by traces of active trypsin degrades zymogen

• The alkaline pH (8.0-9.5) and low CaThe alkaline pH (8.0-9.5) and low Ca2+2+ concentration in concentration in pancreatic secretions promote the degradation rather than pancreatic secretions promote the degradation rather than the activation of trypsinogenthe activation of trypsinogen

• In acute pancreatitis activated trypsin and other enzymes In acute pancreatitis activated trypsin and other enzymes are present in the ducts of the pancreas are present in the ducts of the pancreas

ElastaseElastase • Elastase, like trypsin and chymotrypsin, is a Elastase, like trypsin and chymotrypsin, is a

serine protease that also hydrolyses amides serine protease that also hydrolyses amides and estersand esters

• Attacking bonds next to small amino acids Attacking bonds next to small amino acids residues such as glycine, alanine and serineresidues such as glycine, alanine and serine

• Two types; Two types; – Elastase I, which is anionic exists in serum both Elastase I, which is anionic exists in serum both

free and as a complex with alpha-1 proteinase free and as a complex with alpha-1 proteinase inhibitor; inhibitor;

– Elastase II which is cationic mainly in the Elastase II which is cationic mainly in the bound form with alpha-1 proteinase inhibitorbound form with alpha-1 proteinase inhibitor

ElastaseElastase

• Clinical Significance:Clinical Significance: – Elastase I is increased in acute and relapsing chronic Elastase I is increased in acute and relapsing chronic

pancreatitispancreatitis

– There is some evidence that this enzyme is considerably There is some evidence that this enzyme is considerably more specific for pancreatitis than is amylase. more specific for pancreatitis than is amylase.

– Elevation of elastase I have also been observed in Elevation of elastase I have also been observed in carcinoma of the pancreas, especially carcinoma of the carcinoma of the pancreas, especially carcinoma of the head of the pancreas.head of the pancreas.

– Elastase II is likewise increased in acute pacreatitisElastase II is likewise increased in acute pacreatitis

• Enteropeptidase:Enteropeptidase:– Is a Is a serine proteaseserine protease enzymeenzyme secreted by intestinal mucusa secreted by intestinal mucusa – Cleaves after Lysine if the Lys is preceded by four Asp and not Cleaves after Lysine if the Lys is preceded by four Asp and not

followed by a Pro.followed by a Pro.

• Carboxypeptidase:Carboxypeptidase:– Carboxypeptidases are usually classified into one of several families Carboxypeptidases are usually classified into one of several families

based on their active site mechanismbased on their active site mechanism• Enzymes that use a metal in the active site are called "metallo-Enzymes that use a metal in the active site are called "metallo-

carboxypeptidases". carboxypeptidases". • Carboxypeptidases that use active site Carboxypeptidases that use active site serineserine residues are called residues are called

"serine carboxypeptidases". "serine carboxypeptidases". • Carboxypeptidases that use an active site Carboxypeptidases that use an active site cysteinecysteine are called are called

"cysteine carboxypeptidase" (or "thiol carboxypeptidases")."cysteine carboxypeptidase" (or "thiol carboxypeptidases").

– By substrate preference:By substrate preference:• In this classification system, carboxypeptidases that have a stronger In this classification system, carboxypeptidases that have a stronger

preference for those amino acids containing aromatic or branched preference for those amino acids containing aromatic or branched hydrocarbon chains are called carboxypeptidase A (A for hydrocarbon chains are called carboxypeptidase A (A for aromatic/aliphatic). aromatic/aliphatic).

• Carboxypeptidases that cleave positively charged amino acids Carboxypeptidases that cleave positively charged amino acids (arginine, lysine) are called carboxypeptidase B (B for basic). (arginine, lysine) are called carboxypeptidase B (B for basic).

Induced conformation (Induced conformation (carboxypeptidase A)carboxypeptidase A)

Brush Border Enzymes of The IntestineBrush Border Enzymes of The Intestine

• Aminopeptidase, Leucine aminopeptidaseAminopeptidase, Leucine aminopeptidase

• Oligopeptidases, degrade small peptides such as Oligopeptidases, degrade small peptides such as tetrapeptidestetrapeptides

• Dipeptidyl aminopeptidases, remove dipeptides from the Dipeptidyl aminopeptidases, remove dipeptides from the N-terminal of proteinsN-terminal of proteins

• Tripeptidases and Dipeptidases in the endothelial cellsTripeptidases and Dipeptidases in the endothelial cells

LipasesLipases

• Human lipase is a glycoprotienHuman lipase is a glycoprotien• Lipases are defined as enzymes that hydrolyze glycerol Lipases are defined as enzymes that hydrolyze glycerol

esters of long-chain fatty acids.esters of long-chain fatty acids. • For full catalytic activity and greatest specificity, the For full catalytic activity and greatest specificity, the

presence of bile salts and a cofactor, called co-lipase is presence of bile salts and a cofactor, called co-lipase is required.required.

• Both lipase and colipase are synthesized in the pancreatic Both lipase and colipase are synthesized in the pancreatic acinar cells and secreted by the pancreas in roughly acinar cells and secreted by the pancreas in roughly equimolar quantities.equimolar quantities.

• Lipase measurements are used exclusively to investigate Lipase measurements are used exclusively to investigate pancreatic disorders, usually pancreatitis pancreatic disorders, usually pancreatitis

Brush Border Enzymes of The IntestineBrush Border Enzymes of The Intestine

• Isomaltase (alpha 1,4-glycosidase)Isomaltase (alpha 1,4-glycosidase)

• GlucoamylaseGlucoamylase

• MaltaseMaltase

• SucraseSucrase

• LactaseLactase

• TrehalaseTrehalase

• Sucrase and isomaltase are synthesized as a single Sucrase and isomaltase are synthesized as a single polypeptide chain inside the cellpolypeptide chain inside the cell

LactaseLactase• Secreted by Brunner's glands of the duodenum and glands of Secreted by Brunner's glands of the duodenum and glands of

Lieberkun, works at pH 5.4-6.0 hydrolyze lactose to glucose and Lieberkun, works at pH 5.4-6.0 hydrolyze lactose to glucose and galactose.galactose.

• Lactase deficiency:Lactase deficiency: – Symptoms of flatulence, abdominal discomfort, bloating, or Symptoms of flatulence, abdominal discomfort, bloating, or

diarrhea diarrhea

• Lactase deficiency of two types:Lactase deficiency of two types:– Congenital Lactase DeficiencyCongenital Lactase Deficiency

• Symptoms occur as soon as milk is takenSymptoms occur as soon as milk is taken• Absent or low intestinal lactase in the neonate, however, cannot Absent or low intestinal lactase in the neonate, however, cannot

be taken as proof of congenital deficiency because lactase is be taken as proof of congenital deficiency because lactase is normally the slowest of the oligosaccharidases to reach normal normally the slowest of the oligosaccharidases to reach normal levels in the newborn’s gutlevels in the newborn’s gut

• An abnormal oral lactose tolerance test obtained a few months An abnormal oral lactose tolerance test obtained a few months after birth could also be due to congenital glucose-galactose after birth could also be due to congenital glucose-galactose intoleranceintolerance

LactaseLactase

• Acquired Lactase DeficiencyAcquired Lactase Deficiency– Lactase activity declines as a child agesLactase activity declines as a child ages– The age at which decline begins is genetically determined in an The age at which decline begins is genetically determined in an

Autosomal recessive fashion and differs among ethnic groupsAutosomal recessive fashion and differs among ethnic groups

• Secondary Lactase IntoleranceSecondary Lactase Intolerance– Occurs as a result of reduced enzyme activity following diffuse Occurs as a result of reduced enzyme activity following diffuse

intestinal damage from;intestinal damage from;– Infections (Giardiasis, bacterial overgrowth, or viruses), Infections (Giardiasis, bacterial overgrowth, or viruses), – Ulcerative colitis, Ulcerative colitis, – Celiac disease, and Celiac disease, and – Tropical sprueTropical sprue

• Testing: Testing: hydrogen breath test is used to measure the amount of hydrogen breath test is used to measure the amount of hydrogen in the breath. hydrogen in the breath.

• Ribonuclease (RNase): Ribonuclease (RNase): digestion of dietary nucleic acidsdigestion of dietary nucleic acids

– Ribonuclease A catalyzes cleavage of the phosphodiester bond Ribonuclease A catalyzes cleavage of the phosphodiester bond between the 5'-ribose of a nucleotide and the phosphate group between the 5'-ribose of a nucleotide and the phosphate group attached to the 3'-ribose of an adjacent pyrimidine nucleotide attached to the 3'-ribose of an adjacent pyrimidine nucleotide forming a 2',3'-cyclic phosphateforming a 2',3'-cyclic phosphate

– Ribonuclease B: is a glycosylated derivative of RNase A. Ribonuclease B: is a glycosylated derivative of RNase A.

• DeDeoxyribonuclease (DNase): oxyribonuclease (DNase): digestion of dietary nucleic aciddigestion of dietary nucleic acid

• Polynucleotidases: Polynucleotidases: nucleic acids into nucleotidesnucleic acids into nucleotides

• Nucleosidases: Nucleosidases: nucleosides to give free nitrogen base plus pentose nucleosides to give free nitrogen base plus pentose phosphatephosphate

• Phosphatases: Phosphatases: remove phosphateremove phosphate

• Phospholipases: Phospholipases: phospholipids to produce glycerol, fatty acids, phospholipids to produce glycerol, fatty acids, phosphoric acid, and bases such as choline phosphoric acid, and bases such as choline

• Cholesterol esterase: Cholesterol esterase: cholesterol and fatty acidscholesterol and fatty acids

• Sucrase-Isomaltase deficiency: flatulence, abdominal Sucrase-Isomaltase deficiency: flatulence, abdominal discomfort, bloating, diarrheadiscomfort, bloating, diarrhea

• Colipase deficiency: steatorrheaColipase deficiency: steatorrhea

• Absorption of undigested polypeptides may cause antigenic Absorption of undigested polypeptides may cause antigenic reactions: celiac diseasereactions: celiac disease

• Monosaccharide Malabsorption: caused by single mutation Monosaccharide Malabsorption: caused by single mutation resulted in a defect in the Naresulted in a defect in the Na++-glucose co-transporter -glucose co-transporter carrier mechanism (SGLT1) carrier mechanism (SGLT1)

Disturbances due to malabsorptionDisturbances due to malabsorption

• Anemia: Iron, Vitamin BAnemia: Iron, Vitamin B1212, Folate, Folate• Edema: Products of protein digestionEdema: Products of protein digestion• Tetany: Calcium, Magnesium, Vitamin DTetany: Calcium, Magnesium, Vitamin D• Osteoporosis: Calcium, Product of protein digestion, Osteoporosis: Calcium, Product of protein digestion,

Vitamin DVitamin D• Milk intolerance: LactoseMilk intolerance: Lactose• Bleeding Bruising: Vitamin KBleeding Bruising: Vitamin K• Steatorrhea (fatty stools): Lipids and fat-soluble vitaminsSteatorrhea (fatty stools): Lipids and fat-soluble vitamins• Hartnup disease (defect in intestinal neutral amino acid Hartnup disease (defect in intestinal neutral amino acid

carrier): Neutral amino acids carrier): Neutral amino acids