Enzymes proline specific peptidase

In addition to phosphotriesterase from P. diminuta (PTE) discussed above, two other types of enzymes were found to exhibit phosphotriesterase activity. Interestingly, both are peptidases - the enzymes which in nature hydrolyse a peptide bond. The first one - organophosphorus acid anhydrolase (OPAA) from Alteromonas sp. JD6.5 - is a proline dipeptidase its original activity is to cleave a dipepfide bond with a prolyl residue at the carboxy terminus. The second one - aminopeptidase P (AMPP) from Escherichia coli - is a proline-specific peptidase that catalyses hydrolysis of N-terminal peptide bonds containing a proline residue. ° ... 

Although any of several combinations of proteases can be used, ideally, one or more non-specific endopeptidases should be used first to convert the protein into many small peptides. These small peptides can then be degraded to amino acids by aminopeptidases and prolidase (hydrolyzes X-Pro bonds). Sometimes, carboxypeptidases are also used. Although leucine aminopeptidase has been used as the amino-peptidase (see Hill and Schmidt 1962), it may be preferable to use aminopeptidase M (Rohm and Haas, supplied by Henley and Co. of N.Y.), since this enzyme removes most residues at acceptable rates. Leucine aminopeptidase removes hydrophobic residues most rapidly, whereas some other residues are removed very slowly. Most procedures should probably include the use of prolidase (Miles) since many aminopeptidases do not cleave X-Pro bonds at appreciable rates. If it is found that proline is not released quantitatively by these procedures, the use of citrus leaf carboxypeptidase C (Rohm and Haas) can be tried after the initial endopeptidase hydrolysis and before the addition of aminopeptidase M and prolidase. Carboxypeptidase C (also yeast carboxypeptidase Y - see Hayashi et al. 1973) hydrolyzes proline bonds (as well as all others), but if proline is at or adjacent to the NH2 terminus of a peptide, it would probably not be released. In all procedures a control consisting of the enzymes only should be run in parallel with the hydrolyzed sample, and corrections should be made for any amino acids found by analysis of the control. suhic / /< > , mi... 

---