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Dipeptidyl Aminopeptidase III of Guinea‐Pig Brain: Specificity for Short Oligopeptide Sequences
Author(s) -
Smyth Maria,
O'Cuinn Gerard
Publication year - 1994
Publication title -
journal of neurochemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.75
H-Index - 229
eISSN - 1471-4159
pISSN - 0022-3042
DOI - 10.1046/j.1471-4159.1994.63041439.x
Subject(s) - angiotensin iii , dipeptide , biochemistry , aminopeptidase , chemistry , pepstatin , peptide , amino acid , enzyme , oligopeptide , gel electrophoresis , hydrolysis , angiotensin ii , leucine , protease , angiotensin receptor , receptor
Abstract: A dipeptidyl aminopeptidase III‐type activity has been purified from the cytoplasm of guinea‐pig brain using arginyl‐arginyl‐7‐amido‐4 methylcoumarin as substrate. The enzyme was purified 754‐fold relative to the crude homogenate and with a 12.7% recovery. The purified enzyme was found to have a relative molecular weight of 85,000 and consists of one polypeptide chain of relative molecular weight 80,000, on the basis of its migration on calibrated sodium dodecyl sulphate‐polyacrylamide gel electrophoresis gel. It is highly sensitive to the presence of chelating agents, sulphydryl reactive agents, and the dipeptide Tyr‐Tyr. Dithiothreitol (1 m M ) reduced activity by 28%, and 36 and 65% inhibition was noted with phenylmethylsulphonyl fluoride and puromycin (both at 1 m M ), respectively. Little or no inhibition was observed with bestatin, bacitracin, captopril, amastatin, and arphamenine B. The purified enzyme released dipeptide moieties from a wide range of peptides including enkephalin sequences and also angiotensin sequences up to the octapeptide angiotensin II. These sequences inhibited the hydrolysis of arginyl‐arginyl‐7‐amido‐4‐methylcoumarin by dipeptidyl aminopeptidase III with K i values in the micromolar range. No hydrolysis was observed with angiotensin I or with peptide sequences containing more than 10 amino acids. No hydrolysis was observed also with peptide sequences containing a Pro residue on either side of the sissile bond. Peptides containing less than four amino acids were not hydrolysed.