Serine exopeptidases

The proteolytic enzymes are classified into endopeptidases and exopeptidases, according to their site of attack in the substrate molecule. The endopeptidases or proteinases cleave peptide bonds inside peptide chains. They recognize and bind to short sections of the substrate s sequence, and then hydrolyze bonds between particular amino acid residues in a relatively specific way (see p. 94). The proteinases are classified according to their reaction mechanism. In serine proteinases, for example (see C), a serine residue in the enzyme is important for catalysis, while in cysteine proteinases, it is a cysteine residue, and so on. 

We have chosen to discuss enzyme modification of proteins in terms of changes in various functional properties. Another approach might have been to consider specific substrates for protease action such as meat and milk, legumes and cereals, and the novel sources of food protein such as leaves and microorganisms ( ). Alternatively, the proteases themselves provide categories for discussion, among which are their source (animals, plants, microorganisms), their type (serine-, sulfhydryl-, and metalloenzymes), and their specificity (endo- and exopeptidases, aromatic, aliphatic, or basic residue bond specificity). See Yamamoto (2) for a review of proteolytic enzymes important to functionality. 

Proteases can be subdivided into two major groups exopeptidases cleaving the peptide bond proximal to the amino or carboxy terminal of the substrate, and endopeptidases cleaving distant from the termini (Rao et al., 1998). According to the functional group at the active site, proteases are further classified into four groups serine proteases, aspartyl proteases, cysteine proteases and metalloproteases. Based on the pH optimal for their functioning, proteolytic enzymes can be characterised as alkaline, neutral or acidic proteases. 

Dietary proteins, with very few exceptions, are not absorbed rather they must be digested into amino acids, or di- and tripeptides. Protein digestion begins in the stomach, where proenzyme pepsinogen is autocatalytically converted to pepsin A. Most proteolysis takes place in the duodenum via enzymes secreted by the pancreas, including trypsinogen, chymotrypsinogen and pro-carboxypeptidase A. These serine and zinc proteases are produced in the form of their respective proenzymes they are both endopeptidase and exopeptidase, and their combined action leads to the production of amino acids, dipeptides and tripeptides. 

In a general sense exopeptidases should be the enzymes of choice for stepwise chain assembly since once formed the internal peptide bonds of the growing chain can no longer be proteolytically cleaved from this type of peptidase. Carbox-ypeptidase exhibit superior properties for the stepwise synthesis, especially, carbox-ypeptidase Y (CPD-Y)12101 or other serine peptidases of this type. In principle, aminopeptidases can also be used starting from the C-terminus. Because under these conditions not only the carboxyl component but also the amine component has a free a-amino function, product isolation is more difficult, particularly, if one component is used in excess. Otherwise, stepwise synthesis from the C-terminus is not a problem in chemical peptide synthesis. 

Chymotrypsin is only one member of the family of serine proteases (enzymes that utilize an active-site serine (Ser, S) to cleave the peptide chain). Trypsin, also an endopeptidase obtained from bovine pancreas, is another member of the same family of serine (Ser, S) proteases. Interestingly, however, trypsin cleaves peptides on the carbon side of lysine (Lys, K) and arginine (Arg, R) groups. The catalytic triad discussed above for chymotrypsin also appears to apply here, while still other peptidases have other conserved units that allow them to catalyze the cleavage of peptides at specific sites. While it is not appropriate to provide an exhaustive list of peptidases that have been found to effect protein cleavage, it is important to be aware that in addition to serine (Ser, S) proteases, there are cysteine (Cys, C) proteases and aspartate (Asp, D) proteases and, in addition to endopeptidases, there are aminopeptidases that act on N-termini (N-terminal exopeptidases) and carboxy-peptidases (carboxyl-terminus exopeptidases) that act at the corresponding carboxylic acid termini. 

Proteases are grossly subdivided into two major groups, such as exopeptidases and endopeptidases, depending on their site of action. Based on the functional group present at the active site, proteases are further classified into four prominent groups, such as serine, aspartic, cysteine, and metal-loproteases (Hartley, 1996). There are a few miscellaneous proteases that do not precisely fit into the standard classification and one of them is ATP-dependent proteases (Menon et al., 1987). The flow sheet for classification of peptide hydrolases is given in Figure 9.3. 

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