Studies of three major proteases associated with guinea pig sperm acrosomes.
The major proteases associated with guinea pig sperm were investigated by using immunological and electrophoretic techniques. Three major proteases were detected following sodium dodecyl sulfate-polyacryl-amide gel electrophoresis in gels containing 0.1% gelatin. These enzymes had molecular weights of 47,000, 34,000, and 32,000 relative to reduced protein standards and 58,500, 40,000, and 37,500 relative to unreduced standards. All three protease activities were present in acid extracts of sperm, detergent extracts of sperm, and the soluble acrosomal components of sperm released following induction of the acrosome reaction with the Ca2+-ionophore A23187. As determined by indirect immunofluorescence, an antibody to purified boar acrosin specifically cross-reacted with the acrosomes of guinea pig sperm. Decreased fluorescence was associated with sperm that had lost their acrosomes. Immunoblot analysis demonstrated that this antibody reacted with the 47,000 Mr protease but not the 32,000 and 34,000 Mr proteases. All three proteases were maximally active in the pH 7.0-8.5 region and were inhibited by classical inhibitors of acrosin activity. During a 3-hour incubation period, MgCl2 (10 mM) inhibited the activities of the 32,000 and 34,000 Mr proteases while the 47,000 Mr protease was stimulated. Although these proteases shared properties that would classify them as trypsin-like proteases, only the 47,000 Mr protease could be definitely classified as a member of the proacrosin-acrosin family based upon cross-reaction with an antibody to purified boar acrosin.
Arboleda
C E
CE
Department of Obstetrics and Gynecology, University of Pennsylvania School of Medicine, Philadelphia 19104-6080.
Gerton
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GL
D000176:Acrosin / Q000032:analysis; D000177:Acrosome / Q000201:enzymology*; D000818:Animals; D002122:Calcium Chloride / Q000494:pharmacology; D003429:Cross Reactions; D004591:Electrophoresis, Polyacrylamide Gel; D005455:Fluorescent Antibody Technique; D006168:Guinea Pigs; D006863:Hydrogen-Ion Concentration; D007118:Immunoassay; D008274:Magnesium / Q000494:pharmacology; D015636:Magnesium Chloride; D008297:Male; D008970:Molecular Weight; D010447:Peptide Hydrolases / Q000032:analysis*; D011480:Protease Inhibitors / Q000494:pharmacology; D013094:Spermatozoa / Q000201:enzymology*
D011480:Protease Inhibitors; D015636:Magnesium Chloride; D010447:Peptide Hydrolases; D000176:Acrosin; D008274:Magnesium; D002122:Calcium Chloride