Enzymes aminopeptidase

The metalloproteases include both exopeptidases (e.g., angiotensin-converting enzyme, aminopeptidase-M, and carboxypeptidase-A) and endopeptidases (e.g.,... 

Although the Edman method is by far the most common method of sequencing peptides from the /V-terminus, some enzymic methods are used occasionally. Several aminopeptidases are available commercially, which differ in their specificities. One aminopeptidase from porcine kidney preferentially releases amino acids such as leucine with hydrophobic side-chains. This enzyme does not release /V-terminal Arg or Lys or any amino acid that is followed by Pro. Another enzyme, aminopeptidase M, which is obtained from the microsomal fraction of porcine kidney cells, is less specific and perhaps more useful. It is advisable to examine aliquots of the hydrolysate at intervals by chromatography to determine the order in which amino acids are being released. 

The potent enzyme (aminopeptidase) inhibitor bestatin 161 provides another perfect bridge between the amino- and the hydroxy-acids. At first sight bestatin appears to be a dipeptide but only one of the components, leucine 163 is a normal a-amino acid. The other half 162 is a P-amino acid and also an a-hydroxyacid and that might give us the idea that malic acid 34, with the same absolute configuration at the secondary alcohol, could provide a chiral pool starting material. 

Although the surface-capping agents are usually considered as trouble in catalytic applications, they can actually participate in catalytic reactions. Kim et al. synthesized Pt nanoparticles (1-2 nm in size) by using an enzyme (aminopeptidase) [57]... 

Several enzymes, none of which are completely specific for the enkephalins, are known to cleave Leu- and Met-enkephalin at various peptide bonds. The main enzymes that degrade enkephalin are 2inc metaHopeptidases. The first enkephalin-degrading enzyme to be identified, an aminopeptidase which cleaves the amino terminal Tyr-Gly bond (179), has been shown to be aminopeptidase-N (APN) (180). It is a cytoplasmic enzyme which is uniformly distributed throughout the brain. The increased analgesic activity of synthetic enkephalins substituted by D-amino acids at position 2, eg,... 

Other interesting examples of proteases that exhibit promiscuous behavior are proline dipeptidase from Alteromonas sp. JD6.5, whose original activity is to cleave a dipeptide bond with a prolyl residue at the carboxy terminus [121, 122] and aminopeptidase P (AMPP) from E. coli, which is a prohne-specific peptidase that catalyzes the hydrolysis of N-terminal peptide bonds containing a proline residue [123, 124]. Both enzymes exhibit phosphotriesterase activity. This means that they are capable of catalyzing the reaction that does not exist in nature. It is of particular importance, since they can hydrolyze unnatural substrates - triesters of phosphoric acid and diesters of phosphonic acids - such as organophosphorus pesticides or organophosphoms warfare agents (Scheme 5.25) [125]. 

There are two main classes of proteolytic digestive enzymes (proteases), with different specificities for the amino acids forming the peptide bond to be hydrolyzed. Endopeptidases hydrolyze peptide bonds between specific amino acids throughout the molecule. They are the first enzymes to act, yielding a larger number of smaller fragments, eg, pepsin in the gastric juice and trypsin, chymotrypsin, and elastase secreted into the small intestine by the pancreas. Exopeptidases catalyze the hydrolysis of peptide bonds, one at a time, fi"om the ends of polypeptides. Carboxypeptidases, secreted in the pancreatic juice, release amino acids from rhe free carboxyl terminal, and aminopeptidases, secreted by the intestinal mucosal cells, release amino acids from the amino terminal. Dipeptides, which are not substrates for exopeptidases, are hydrolyzed in the brush border of intestinal mucosal cells by dipeptidases. 

The proteases are secreted as inactive zymogens the active site of the enzyme is masked by a small region of its peptide chain, which is removed by hydrolysis of a specific peptide bond. Pepsinogen is activated to pepsin by gastric acid and by activated pepsin (autocatalysis). In the small intestine, trypsinogen, the precursor of trypsin, is activated by enteropeptidase, which is secreted by the duodenal epithelial cells trypsin can then activate chymotrypsinogen to chymotrypsin, proelas-tase to elastase, procarboxypeptidase to carboxypepti-dase, and proaminopeptidase to aminopeptidase. 

Binding of bidentate sulfur ligands can also be of importance in the inhibition of zinc containing enzymes. An aminopeptidase which is essential for vancomycin antibiotic resistance in... 

Aminopeptidase A is another brush border membrane enzyme which has been the subject of various studies [79,81,83-86], It has been found in the intestinal brush border membrane of humans, rabbits, rats, and pigs and is active against peptides with acidic amino acids at the amino terminus. Its activity against dipeptides is more limited. Shoaf et al., isolated three rat brush border aminopeptidases with distinct but somewhat overlapping substrate specificities. These enzymes had preference for dipeptides containing methionine, arginine, or aspartic acid and glycine. The optimal pH for activity of aminopeptidase was reported to be 7-8. 

Dipeptidyl aminopeptidase IV hydrolyzes substrates with free a-amino groups. Peptide bonds involving the carboxy group of either Pro or Ala are cleaved by this enzyme to X-Pro or X-Ala, where X may be any amino acid. It has been shown that peptides with the X-Pro moiety are hydrolyzed more completely than those with X-Ala [79],... 

Graham, M., Smith, T.S., Munn, E.A. and Newton, S.E. (1993) Recombinant DNA molecules encoding aminopeptidase enzymes and their use in the preparation of vaccines against helminth infections. Patent No. WO 93/23542. 

In addition to oxidative and conjugative metabolism, other enzymes are also present in the Caco-2 model, though at lower levels than in the human enterocyte in vivo. These are mainly apical cell surface peptidases, such as aminopeptidases... 

Bestatin (ubenimex) is a potent inhibitor of aminopeptidase N and aminopeptidase B,89 which was isolated from a culture filtrate of Streptomyces olivoreticuli during the search for specific inhibitors of enzymes present on the membrane of eukaryotic cells.90 Inhibitors of aminopeptidase activity are associated with macrophage activation and differentiation, Bestatin has shown significant therapeutic effects in several clinical trials.91 In a multi-institutional study,92 patients with acute non-lymphocytic leukemia (ANLL) were randomized to receive either Bestatin or placebo orally after completion of induction and consolidation therapy, and concomitant with maintenance chemotherapy. Remission duration was prolonged in the Bestatin group, although this difference did... 

Aminopeptidase N. This enzyme works in a manner similar to carboxypeptidase Y, but cleaves amino acids from the N-terminus. It can be used in the same way as carboxypeptidase Y. 

It was clear for some time that a number of zinc enzymes required two or more metal ions for full activity, but in the absence of X-ray structural data the location of these metal centres with regard to one another was often uncertain. When the first 3-D structures began to appear, it became clear that the metals were in close proximity. A particular feature of many of these enzymes was the presence of a bridging ligand between two of the metal sites, usually an Asp residue of the protein, which is occasionally replaced by a water molecule. While some of the sites contain only Zn ions, several contain Zn in combination with Cu (in cytosolic superoxide dismutases) Fe (in purple acid phosphatases) or Mg (in alkaline phosphatase and the aminopeptidase of lens). 

Non-corrin cobalt has a number of interesting applications in the chemical industry, for example in the hydroformylation (OXO) reaction between CO, H2 and olefins. A number of non-corrin Co-containing enzymes have been described, including methionine aminopep-tidase, prolidase, nitrile hydratase and glucose isomerase. We describe the best characterized of these, namely the E. coli methionine aminopeptidase, a ubiquitous enzyme, which cleaves N-terminal methionine from newly translated polypeptide chains. The active site of the enzyme (Figure 15.13) contains two Co(II) ions that are coordinated by the side-chain atoms of five amino acid residues. The distance between the two Co2+ is similar to that between the two Zn2+ atoms in leucine aminopeptidase, and indeed the catalytic mechanism of methionine aminopeptidase shares many features with other metalloproteases, in particular leucine aminopeptidases. 

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