Properties of a proteolytic enzyme from rabbit sperm acrosomes.

Abstract

Acrosin is a proteolytic enzyme extractable from sperm acrosomes and essential for penetration of the zona pellucida of the ovum by spermatozoa. It has properties in common with both trypsin and plasmin having an optimum pH of 8.0, hydrolyzing benzoyl arginme ethyl ester (BAEE) and tosyl arginine methyl ester (TAME) but not benzoyl tyrosine ethyl ester (BTEE), or acetyl tyrosine ethyl ester (ATEE), being stimulated by calcium ions and stable at pH 3.0. Acrosin differs from pancreatic trypsin in that it hydrolyzes BAEE more rapidly than TAME, is less effectively inhibited by ovomucoid and pancreatic trypsin inhibitor, has a higher molecular weight, undergoes rapid loss of activity even in the presence of calcium and produces fewer peptides on incubation with oxidized nbonuclease. One of two trypsin inhibitors isolated from guinea pig seminal vesicles inhibits acrosin and trypsin but not plasmin, whereas the other inhibits all three enzymes. Acrosin further differs from plasmin in that it has a different molecular weight and is more unstable. Inhibition by diisopropyl fluorophosphate and tosyl lysine chloromethyl ketone shows a similarity to trypsins and lack of inhibition by tosyl phenylalanine chloromethyl ketone indicates that acrosin is not chymotrypsin-like. The enzyme is also inhibited by soybean trypsin inhibitor. Acrosin appears to be a unique proteolytic enzyme and therefore deserves a specific name. Previously we described the purification of an acrosomal trypsin-like enzyme, acrosin, from epididymal sperm with a molecular weight of 55,000 (Polakoski et a!., 1971). The presence of such a proteolytic enzyme in sperm was first reported by Yamane (1935a, b) and shown to be trypsin-like by Buruiana (1956). Hartree and Srivastava (1965), and Srivastava, Adams, and Hartree (1965) showed that acrosomal extracts prepared from ram sperm removed the zona pellucida from rabbit ova in vitro. Stambaugh and Buckley (1968, 1969) confirmed these results using similar extracts prepared from epididymal rabbit sperm. These authors showed that the protease demonstrated to be localized in acrosomal extracts by Srivastava ci a!. (1965) was trypsin-like in nature and responsible for the removal of the zona. It is most likely the same enzyme studied by Buruiana (1956). In vitro fertilization was inhibited by soybean trypsin inhibitor (Stambaugh ci a!., 1969). Fertilization in vivo was inhibited by preincubation of capacitated sperm with pancreatic tryp