Guinea pig testicular proacrosin-acrosin system: further characterization of the active enzyme.
In this paper, the characteristics of a highly stable, 34,000 molecular weight form of guinea pig (GP) acrosin are compared with those of acrosins from other mammalian species. GP acrosin, like acrosins from other species, is stable at pH 3.0, has a pH optimum of 8.0, and is inhibited by natural trypsin inhibitors and N-alpha-p-tosyl-L-lysine chloromethyl ketone. Its lack of inhibition by tosyl-phenylalanine chloromethyl ketone indicates that it has a specificity similar to trypsin but not chymotrypsin. The activity of GP acrosin was stimulated by Ca2+ below 75 mM. The enzyme was markedly inhibited by Hg2+, but only weakly inhibited by other metal cations. The disulfide reductants dithiothreitol and 2-mercaptoethanol both inhibited GP acrosin, as did the sulfhydryl reactant, iodoacetic acid. The Michaelis-Menten constant for GP testicular acrosin-catalyzed hydrolysis of the N-benzyloxycarbonyl-L-arginyl amide of 7-amino-4-trifluoromethylcoumarin at pH 8.0 was calculated from Lineweaver-Burk plots to give a value of Km = 2.0 x 10(-5) M with Vmax = 500 mumoles/min/mg protein. The corresponding lysine substrate, the N-benzyloxy-carbonyl L-lysine amide of 7-amino-4-trifluoromethyl-coumarin, had a higher Km = 4.6 x 10(-5) M and lower Vmax = 135 mumoles/min/mg protein, in accord with the substrate preference seen with other mammalian acrosins.(ABSTRACT TRUNCATED AT 250 WORDS)
Adekunle
A O
AO
Department of Obstetrics and Gynecology, University of Pennsylvania School of Medicine, Philadelphia 19104-6080.
Storey
B T
BT
Teuscher
C
C
D000176:Acrosin / Q000378:metabolism*; D000818:Animals; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004792:Enzyme Precursors / Q000378:metabolism*; D006168:Guinea Pigs / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008297:Male; D012044:Regression Analysis; D012697:Serine Endopeptidases / Q000378:metabolism*; D013045:Species Specificity; D013379:Substrate Specificity; D013737:Testis / Q000201:enzymology*
D004791:Enzyme Inhibitors; D004792:Enzyme Precursors; D012697:Serine Endopeptidases; C022438:proacrosin; D000176:Acrosin