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J. Biol. Chem., Vol. 264, Issue 23, 13586-13590, Aug, 1989
TD Noland, LS Davis and GE Olson
Following sodium dodecyl sulfate-polyacrylamide gel electrophoresis,
proacrosin has been identified in extracts of intact guinea spermatozoa as
a major silver staining band which reacted immunologically with antibodies
made against purified proacrosin from guinea pig testis. Proacrosin
exhibited an approximate Mr of 50,000 and was rapidly converted to an Mr
45,000 protein following induction of the acrosome reaction with 2.0 mM
CaCl2 and 1 micrograms/ml A23187. Apical segments isolated at pH 6.0 from
guinea pig spermatozoa also contained a major silver staining band of Mr
50,000 which cross-reacted with antibodies to guinea pig testis proacrosin.
Subcellular fractionation of spermatozoa indicated that proacrosin remained
in the particulate fraction of homogenized spermatozoa and was enriched
within the isolated acrosomal apical segment. When apical segments isolated
at pH 6.0 were incubated at pH 7.5, proacrosin was rapidly converted to the
Mr 45,000 form observed in spermatozoa undergoing the acrosome reaction.
The conversion process in isolated apical segments was inhibited by
leupeptin and was accelerated in the presence of calcium, magnesium, and
manganese. Zinc completely inhibited the conversion of proacrosin to the Mr
45,000 protein. Neither proacrosin nor the Mr 45,000 protein were released
into the supernatant fluid during the incubation of apical segments at pH
7.5. Furthermore, the proteins were resistant to solubilization by 150 mM
NaCl and 1% Triton X-100 but were solubilized by treatment of apical
segments with 1 M NaCl. These results provide evidence as to the identity
and subcellular distribution of proacrosin in intact guinea pig sperm prior
to zymogen conversion and suggest that isolated apical segments exhibit a
subset of the exocytotic reactions leading to completion of the acrosome
reaction.
Regulation of proacrosin conversion in isolated guinea pig sperm acrosomal apical segments
Department of Cell Biology, Vanderbilt University School of Medicine, Nashville, Tennessee 37232.
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