Evidence for a calsequestrin-like calcium-binding protein in human spermatozoa.
Human spermatozoa were investigated for the presence of protein(s) recognized by antibodies against calsequestrin, the high capacity, moderate affinity Ca2(+)-binding protein, originally described in striated muscle fibers. Western immunoblots of detergent-soluble sperm extracts probed with polyclonal antibodies raised against human skeletal muscle calsequestrin identified a strongly cross-reactive protein. This protein resembles muscle calsequestrin in many respects. In fact, its migration in sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) is pH dependent, its apparent molecular mass being 64 kDa in alkaline SDS-PAGE and 44 kDa in neutral SDS-PAGE; its isoelectric point is acidic (4.6); it is metachromatically stained blue by the carboxycyanine dye, Stains-All; it is a Ca2(+)-binding protein (45Ca blot overlay). Indirect immunofluorescence experiments showed that the immunoreactive protein has an intracellular localization confined to the tail mid-piece. From these findings we conclude that human sperm cells express a protein structurally and antigenically related to skeletal muscle calsequestrin; a basis for a novel interpretation of Ca2(+)-mediated events in spermatozoa is thus provided.
Department of Biology, University of Milano, Italy.
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Berruti
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Porzio
D015153:Blotting, Western; D002135:Calcium-Binding Proteins / Q000032:analysis*; D002155:Calsequestrin / Q000032:analysis* / Q000276:immunology; D003429:Cross Reactions; D015180:Electrophoresis, Gel, Two-Dimensional; D005455:Fluorescent Antibody Technique; D006801:Humans; D008297:Male; D009124:Muscle Proteins / Q000032:analysis*; D013094:Spermatozoa / Q000032:analysis*
D002135:Calcium-Binding Proteins; D002155:Calsequestrin; D009124:Muscle Proteins