Purification,crystallisation and characterisation of carboxypeptidase from wheat bran

A carboxypeptidase was purified and crystallised from wheat bran. Disc gel electrophoresis at pH 4·0 and ultracentrifugal analysis revealed that the enzyme was essentially homogeneous. The sedimentation constant and isoelectric point were determined to be 6·3 S and 6·0, respectively. The molecular weight of the enzyme was estimated to be 118,000 by a gel filtration method. The enzyme liberated carboxyl terminal amino acid residues from a wide range of N-substituted dipeptides and tripeptides which contain l-proline. It had a pH optimum at pH 4·0 for Z-Glu-Tyr (Z-benzyloxycarbonyl). The K_m and k_cat values for Z-Glu-Tyr at pH 4·0 and 30°C were 0·19 mm and 20 s^?1, respectively. The enzyme hydrolysed Z-Gly-Pro-Leu-Gly-Pro and bradykinin sequentially at pH 4·0 from their carboxyl terminal amino acid residues. The enzyme activity was completely inhibited by DFP.