Abstract
A proacrosin conversion inhibitor present in boar spermatozoa has been purified and initially characterized. Purification methods included sequential acid extractions of washed spermatozoa at pH 4.0, pH 3.5, and pH 2.5 followed by successive gel filtrations of the pH 2.5 sperm extract supernatant over Sephadex G-75 and G-50. The resulting 8.8-fold purified materials were judged to be homogeneous by sodium dodecyl sulfate-polyacrylamide disc gel electrophoresis, had an estimated molecular weight of 12,800, and a constant specific activity of 65 units/mg. Treatment with the proteinases acrosin, trypsin, or chymotrypsin destroyed the highly purified proacrosin conversion inhibitor, indicating that it is a protein. Additional properties of the inhibitor included stability to long periods of storage at pH 3.0 and 4 degrees C, stability to boiling and lyophilization, and an absolute requirement for divalent cations to maintain activity. The highly purified proacrosin conversion inhibitor does not inhibit acrosin. Therefore, it apparently acts to prevent proacrosin conversion by selectively inhibiting the zymogen's self-catalyzed conversion mechanism.
Highlights
PURIFICATION AND INITIAL CHARACTERIZATION OF A BOAR SPERM PROTEIN WHICH PREVENTS THE CONVERSION OF PROACROSIN INTO ACROSIN*
Purification methods included sequential aciedx- molecular weight factor in guinea pig spermatozoa appeared tractions of washed spermatozoa atpH 4.0, pH 3.5, and toregulate proacrosinconversion into acrosinprior toor pH 2.5 followed by successive gel filtrations of thepH 2.5 sperm extract supernatant over SephadeGx-75 and G-50
Analytical lization, and an absolute requirement for divalentcat- disc gel electrophoresis reagents and equipment were obtained from ions to maintainactivity
Summary
PURIFICATION AND INITIAL CHARACTERIZATION OF A BOAR SPERM PROTEIN WHICH PREVENTS THE CONVERSION OF PROACROSIN INTO ACROSIN*. A proacrosin conversion inhibitor present in boar act to preventproacrosin conversion into acrosin. Spermatozoa has been purified and initially character- Goodpasture et al [8] have recently reported that a low ized. Purification methods included sequential aciedx- molecular weight factor in guinea pig spermatozoa appeared tractions of washed spermatozoa atpH 4.0, pH 3.5, and toregulate proacrosinconversion into acrosinprior toor pH 2.5 followed by successive gel filtrations of thepH 2.5 sperm extract supernatant over SephadeGx-75 and G-50. Treatment with the produringsperm“capacitation”andthe “acrosomereaction.” Earlier findings reported by this laboratory demonstrated that boar sperm extracts contained a similar factor(s) [9]. In this report,the purification andinitialcharacterization of the proacrosin conversion inhibitor found in boar spermatozoais described
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