Transcript Digestive enzymes

Digestive enzymes
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Various organs in digestion and
absorption
 Pancreas is the
major organ that
synthesizes the
digestive enzymes
Small intestine is a principal site
of digestion and absorption
…and there are 3 compartments where digestion and absorption occur:
 Pancreatic enzymes together with bile are poured into the lumen of the
descending part of the duodenum
 Digestion of oligomers of AA and saccharides is accomplished by the
enzymes in the luminal plasma membranes of enterocytes; these
enzymes – usually glycoproteins
 Hydrolysis of di- and tripeptides occurs in the cytoplasm of enterocytes
Zymogens
 Digestive enzymes are usually synthesized as larger inactive precursors
– zymogens
 Otherwise they would digest the tissues that synthesize them:
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acute pancreatitis: premature activation of digestive enzymes
produced by pancreas → auto-digestion of pancreas; activated
phospholipase A2 converts lecithin to lysolecithin that can damage
cell membranes
Synthesis of zymogens
 Proteins destined for secretion are
synthesized on polysomes of the RER
 Their N-terminus contains a signal sequence
→ release of the protein into ER; then, the
signal sequence may be clipped off
 Transport to the Golgi complex
 The proteins are stored in vesicles; after
stimulus, granules move to the luminal
plasma membrane (PM) and fuse with PM
…exocytosis
Zymogens are activated by
proteolysis
 Proenzymes (zymogens) are activated by proteolytic cleavage in the
lumen of the GIT:
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pepsinogen
trypsinogen
chymotrypsinogen
proelastase
procarboxypeptidases
prophospholipases
Activation of pepsinogen
 Pepsinogen is secreted from the stomach cells
 Pepsinogen is activated by the proteolytic removal of 44 AA from its
N-terminus – either as an intramolecular reaction or by active pepsin
 This reaction takes place at pH values below 5
Activation of pancreatic zymogens
in the lumen of the small intestine
chymotrypsinogen, proelastase,
procarboxypeptidases, prophospholipase
enteropeptidase
(produced in duodenum)
trypsinogen
trypsin
– 6 N-terminal AA
autocatalytic activation
chymotrypsin, elastase,
carboxypeptidases, phospholipase
„Strategies“ that prevent
premature zymogen activation
 At pH>2, the peptide (44 AA) clipped of pepsinogen remains bound to
pepsin, masking its active site; it is released by a drop of pH below 2 or
by further degradation by pepsin
 Pancreatic secretory trypsin inhibitor (PSTI), a small polypeptide, blocks
any trypsin that is erroneously activated within the pancreas
Regulation of secretion
 Through secretagogues that interact with the receptors on the surface of
the exocrine cells → signal cascade leading to fusion of granules with PM
Organ
Secretion
Secretagogue
Salivary gland
NaCl, amylase
Stomach
HCl, pepsinogen
acetylcholine, histamine,
gastrin (peptide)
Pancreas
NaCl, enzymes
acetylcholine, cholecystokinin
NaHCO3, NaCl
secretin
acetylcholine
 Cholecystokinin: peptide secreted by cells of small int. after stimulation by AA and
peptides from gastric proteolysis, by FA, and by acid pH
 Secretin: peptide secreted by cells of small int.; stimulated by luminal pH < 5
DIGESTION OF PROTEINS
 By peptidases (proteases):
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endopeptidases – attack internal bonds:
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pepsin
trypsin
chymotrypsin
elastase
exopeptidases – cleave off 1 AA at a time from the:
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C-terminus – carboxypeptidases
N-terminus – aminopeptidases
Classes of peptidases
Type
Active site
pH optimum
Serine proteases
Ser, His, Asp
7-9
Cysteine proteases
Cys, His
3-6
Aspartate proteases
2 x Asp
2-5
Metalloproteases
Zn2+ (coordinated to AA)
7-9
Peptidases hydrolyze the peptide bond
…and differ in substrate specificity:
Pepsins
 Acid in the stomach serves to kill off microorganisms and to denature
proteins (denaturation makes proteins more susceptible to proteolysis)
 Pepsins are acid stable and pH optimum is about 2!!!
 Major products of pepsin action: larger peptide fragments and some free
AA; this mix = peptone
 Importance lies mainly in generation of peptides and AAs that stimulate
cholecystokinin release in the duodenum
Pancreatic enzymes
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trypsin
chymotrypsin
elastase
carboxypeptidases
 Active at neutral pH  depend on neutralization of gastric HCl by
pancreatic NaHCO3
 The combined action of pancreatic peptidases results in the formation
of free AA and small peptides (2-8 AA)
Intestinal peptidases
 Luminal surface of intestinal epithelial cells contains endopeptidases,
aminopeptidases, and dipeptidases that cleave oligopeptides released by
pancreatic peptidases
 Products: AA, di- and tripeptides → absorbed by enterocytes
 Di- and tripeptides are hydrolyzed by intestinal cytoplasmic peptidases
 AA are absorbed into the portal blood
DIGESTION OF SACCHARIDES
 1) Polysaccharides (starch, glycogen) are attacked by -amylase, which is
present in saliva and pancreatic juice (more important)
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-amylase attacks the internal -1,4-glucosidic bonds  products:
maltose, maltotriose, -limit dextrins
 2) Hydrolysis of oligosaccharides is carried out by surface enzymes of the
intestinal epithelial cells – disaccharidases and oligosaccharidases
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These enzymes – often exoglycosidases
Saccharide absorption
 End products: monosaccharides, mainly D-glucose, D-galactose,
D-fructose
 These are transported by a carrier-mediated process into enterocytes and
then into the blood of the portal venous system
Not everything can be digested
 Many plant polymers, including celluloses, hemicelluloses, inulin,
pectin, are resistant to human digestive enzymes
 A small percentage of this „dietary fibre“ is hydrolyzed and then
anaerobically metabolized by the bacteria of the lower intestinal tract
 This bacterial fermentation produces H2, CH4, CO2, H2S, acetate,
propionate, butyrate, lactate
Lactase deficiency
 Experienced as milk intolerance
 Cause:
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a) genetic defect
b) decline of lactase activity with age
c) decline of activity due to an intestinal disease
 Inability to absorb lactose  accumulation and bacterial fermentation of
lactose  production of gas (distension of gut, flatulence); osmotically
active solutes draw water into the intestinal lumen (diarrhea)
Lysozyme
 Hydrolyzes -1,4-glycosidic bonds in the bacterial cell wall polysaccharide
peptidoglycan
 Kills only some types of bacteria
DIGESTION OF LIPIDS
 Lipids – sparingly or not at all soluble in aqueous solutions
 Two problems have to be overcome:
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poor accessibility of the substrate to the enzyme
aggregation of products of hydrolysis to larger complexes that
are hard to absorb
Steps in lipid digestion & absorption
Lipid digestion is initiated in stomach
 In the stomach, acid-stable lipase, secreted by stomach (gastric lipase)
and by lingual glands (lingual lipase), converts TG mostly into FA and
1,2-diacylglycerols (small amount of monoAG is also produced)
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The products possess both polar and non-polar groups  act as
surfactants: stabilize the water-lipid interface  dispersion of the lipid
phase into smaller droplets (emulsification)  better availability of the
substrate to the lipases.
 These lipases have the unique ability to initiate the degradation of
maternal milk fat globules
Pancreatic lipase
 Cleaves acylglycerols mainly to FA and
2-monoacylglycerols
 Requires solubilization of the substrate
 Also requires colipase (secreted by the
pancreas) that anchors and activates the
enzyme
 Absorption of resulting FA and monoAG
requires bile salts micelles
Digestion of phospholipids
 By phospholipases, especially by phospholipase A2 (requires bile acids
for activity):
 FA and lysophospholipids are absorbed from the bile acid micelles
 In the intestinal mucosa, the absorbed lysophospholipids are reacylated
with acyl-CoA
Hydrolysis of cholesterol esters
 By pancreatic cholesterol esterase
 The free cholesterol is transported in the bile acid micelles and
absorbed through the brush border
 Here, it is reacylated with acyl-CoA
Bile acid micelles solubilize lipids
 Primary bile acids are synthesized by the liver and in peroxisomes, they
are conjugated with glycine or taurine (H2N-CH2CH2SO3-)
 A portion of the primary bile acids is subjected to the modifications by
intestinal bacteria → secondary bile acids
 Primary and secondary bile acids are reabsorbed by the ileum into the
portal blood, taken up by the liver, and then resecreted into the bile
…enterohepatic circulation
Bile acid has a hydrophobic surface and
a hydrophilic surface
 The most abundant bile salt in humans – glycocholate:
Bile acid micelles
 Hydrophobic region of the bile salt is
oriented from the water molecules x
hydrophilic region interacts with
water
 Mixed micelles contain (beside bile
acids) phospholipids and cholesterol,
or FA and acylglycerols; FA and
phospholipids form a bilayer in the
interior, bile salts occupy the edge.
 Released FA and monoacylglycerols are incorporated into bile acids
micelles
 Micelles move lipids from the intestinal lumen to the cell surface where
absorption occurs
 Micelles also serve as transport vehicles for vitamins A, K
 Fat malabsorption can result from pancreatic failure or lack of bile acids
 bulk of unabsorbed lipids is excreted with the stool…steatorrhea
Fat digestion and absorption
Most absorbed lipids are
incorporated into chylomicrons
 Within the intestinal cell (after absorption):
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FA of medium chain lenght (6-10C) pass into the portal blood without
modification
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long-chain FA (> 12C) are bound to a fatty acid binding protein in the
cytoplasm and transported to ER, where they are resynthesized to TG
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TG form lipid globules to which phospholipids, cholesterol (esters),
and apolipoproteins adsorb – chylomicrons
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chylomicrons migrate through the Golgi to the basolateral
membrane, they are released, and pass into the lymphatics
DIGESTION OF NUCLEIC ACIDS
 Pancreatic enzymes hydrolyze dietary nucleic acids:
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ribonucleases
deoxyribonucleases
endo- as well as exonucleases
 Polynucleotidases of the small intestine complete the hydrolysis to
nucleotides which are then hydrolyzed to nucleosides by phosphatases
and nucleotidases
 Nucleosides are used as such or undergo degradation by nucleosidases
/ nucleoside phosphorylases to free bases and pentose-1-phosphate
 Purine nucleosides are:
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A) catabolized to uric acid
B) alternatively, purines are released and used for resynthesis of NA
 Pyrimidine nucleosides are:
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A) catabolized to NH4+, CO2,
and β-aminoisobutyrate or
β-alanine, respectively, that
are partially converted to
(methyl)malonyl-CoA
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B) absorbed intact and
utilized for the resynthesis
of nucleic acids