Identification and preliminary characterization of two distinct bovine seminal Pz-peptidases.
Two peptidases hydrolyzing the Pz-peptide substrate were identified in bovine semen. Each Pz-peptidase was strongly inhibited by chelating agents, suggesting both were metallopeptidases. However, these peptidases could be distinguished by other properties and were designated Pz-peptidases A and B. Pz-peptidase A hydrolyzed the Pz-peptide at the Leu-Gly bond, was inhibited by tosylphenylethylchloromethylketone (TPCK) but not by phosphoramidon and had a pH optimum near 6, whereas Pz-peptidase B cleaved the Pro-Leu bond, was inhibited by phosphoramidon but not by TPCK and had a pH optimum near 7. Seminal plasma, light particulates and cytoplasmic droplets contained almost exclusively Pz-peptidase A, and Pz-peptidase A predominated in sperm extracts. Pz-peptidase B was found primarily in sperm extracts, but Pz-peptidase B activity was also present in ultralight particulates. Pz-peptidase A of spermatozoa required Triton X-100 for complete extraction, but Pz-peptidase B was solubilized from spermatozoa by nitrogen decompression without detergents. Pz-peptidase B was inhibited by several detergents. In particular, addition of 0.1% Hyamine 2389 to sperm extracts inhibited 99% of the Pz-peptidase B activity. Thus, Pz-peptidase B may have been overlooked in previous studies employing extraction of spermatozoa with Hyamine 2389. The properties of both seminal PZ-peptidases were different from those of purified bovine testicular PZ-peptidase, suggesting that PZ-peptidases from these sources were not identical.
D000818:Animals; D002417:Cattle; D010450:Endopeptidases / Q000302:isolation & purification* / Q000378:metabolism; D006020:Glycopeptides / Q000494:pharmacology; D006863:Hydrogen-Ion Concentration; D008297:Male; D008666:Metalloendopeptidases; D009842:Oligopeptides; D011480:Protease Inhibitors; D012661:Semen / Q000201:enzymology*; D013094:Spermatozoa / Q000201:enzymology; D013379:Substrate Specificity; D014108:Tosylphenylalanyl Chloromethyl Ketone / Q000494:pharmacology
D006020:Glycopeptides; D009842:Oligopeptides; D011480:Protease Inhibitors; C043091:PZ-peptide; D014108:Tosylphenylalanyl Chloromethyl Ketone; D010450:Endopeptidases; D008666:Metalloendopeptidases; C022510:thimet oligopeptidase; C008890:phosphoramidon