Carboxypeptidase, pancreatic inhibitors

Competitive inhibition of the carboxypeptidase from A. saitoi by small substrates was found with hydrocinnamic acid, indole-3-propionic acid, and 4-phenylbutyric acid [80], The K for hydrocinnamic acid inhibition was 4 x 10 4 M. Diisopropylfluorophosphate (DFP) and tosyl-L-phenylalanylchloromethane (TPCK) were also powerful inhibitors of the carboxypeptidase from A. oryzae (80). />-Chloromercuribenzoate (PCMB) and iodoacetic acid were also powerful inhibitors of the carboxypeptidase from A. saitoi, while the inhibitors of DFP, TPCK, PCMB, and iodoacetic acid on the carboxypeptidase from A. saitoi were less than that of A. oryzae [80], As the carboxypeptidase activity of A. saitoi has no effect when used with ethylenediaminetetraacetate (EDTA) and o-phenanthroline, the enzyme is a different type of carboxypeptidase from those of the pancreatic sources, carboxypeptidase A and carboxypeptidase B [80],... 

The discovery of teprotide led to a search for new, specific, orally active ACE inhibitors. Ondetti et al. (172) proposed a hypothetical model of the active site of ACE, based on analogy with pancreatic carboxypeptidase A, and used it to predict and design compounds that would occupy the carboxy-terminal binding site of the enzyme. Carboxyalkanoyland mer-captoalkanoyl derivatives of proline were found to act as potent, specific inhibitors of ACE and 2-D-methyl-3-mercaptopropanoyl-L-proline (131) (captopril) was developed and launched in 1981 as an orally active treatment for patients with severe or advanced hypertension. Captopril, modeled on the biologically active peptides found in the venom of the pit viper, made an important contribution to the understanding of hypertension and paved the... 

Metalloprotease inhibitors - also known as metalloproteases or zinc proteases - are proteolytic enzymes of which the activity depends on metal ions, normally bound Zn -. Examples of metalloproteases are the pancreatic enzymes carboxypeptidase A and B, elastase, the well-characterized bacterial enzyme thermolysin and the collagenase family (found in both bacterial and mammalian cells, fibroblast collagenase, neutrophil elastase, gelatinase). 

EXAMPLE 5.15 X-ray analysis of crystals of the pancreatic exopeptidase carboxypeptidase A, with a bound pseudo substrate (z. false substrate that is not degraded by the enzyme, i.e., an inhibitor) indicates that the susceptible peptide bond is twisted out of the normal planar configuration that is usually seen with peptide bonds (Chap. 4). This distortion leads to a loss of resonance energy in the bond, and enhances its susceptibility to hydrolytic attack. 

Pancreatic enzymes a group of at least 12 digestive enzymes, including some of the most investigated of all enzymes Autolysis of the pancreas does not occur, because the proteolytic enzymes, trypsin, chymo-trypsin A and B, elastase and carboxypeptidase A and B, and phospholipase A2 are synthesized and stored in the pancreas as inactive zymogens. The other P. e. require effectors for optimal activity, which are present in the duodenum. Trypsin inhibitors in the pancreatic tissue and secretion afford additional protection against proteolytic destruction by active P.e. With the exception of cholesterol esterase (M, 400,000), the M, of P.e. lie between 13,700 (ribonu-clease) and 50,000 (a-amylase). 

Supplementing chick diets with a protein fraction, isolated from raw Russet Burbank potato tubers, enriched in proteinase inhibitors, had previously been shown to severely depress their growth (5). The inhibitor-rich fraction contained at least six well characterized inhibitors of mammalian pancreatic digestive proteinases trypsin, chymotrypsin, elastase and carboxypeptidases A and B (5). Since the fraction contained an array of proteinase inhibitors it was not known if CPI contributed to the growth depressing activities. 

Trypsin inhibitors in animal diets are known to cause pancreatic hypertrophy and hyperplasia. We therefore monitored pancreas weight and enzymic contents in all of the above experiments. The pancreata were removed from the chicks at the termination of the experiments, weighed, and assayed for activities of trypsin, chymotrypsin and carboxypeptidases A and B, after activating their zymogens with either trypsin or enterokinase. 

Figure 4.2. Initial velocity versus substrate concentration plot for pancreatic carboxypeptidase A in the absence and presence of the reversible inhibitor /S-phenylpropionate.

Fig. 3.13. Proportionality of the loss of accessible surface area to the molecular weight of proteins (from Janin, 1976). Different proteins insulin, rubredoxin, pancreatic trypsin inhibitor, HIPIP, calcium binding protein, ribonuclease S, lysozyme, staphylococcal nuclease, papain, chymotrypsin, concanavalin A, subtilisin, thermolysin, carboxypeptidase A.

The formation of tertiary structure buries a certain amount of secondary structure surfaces which varies from 32% to 60%. The proportion of the surface which is buried varies for each protein and increases with the molecular weight of the protein. In fact, it is the proportion of the nonpolar surface that becomes buried during folding, which increases with the molecular weight. From 60% for pancreatic trypsin inhibitor, it varies up to 79% in carboxypeptidase among the six proteins considered by Chothia... 

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