Binding of a bovine oviductal fluid catalase to mammalian spermatozoa.
Hydrogen peroxide (H2O2) is a reactive oxygen species that at low concentration is toxic to sperm. H2O2 inhibits not only sperm viability but also the acrosome reaction, sperm-egg binding, and oocyte penetration. Catalase activates the decomposition of H2O2 into water and oxygen, thus removing an initiator of free radical chain reactions leading to lipid peroxidation. Since the oviduct is known to enhance sperm survival, we hypothesized that it might secrete catalase. We found that oviductal fluid, harvested from washed cells collected at the slaughterhouse, possessed catalase-specific activity that varied during the estrous cycle. Catalase activity increased during the cycle and reached its maximal level just before ovulation (Days 18-20). No significant difference in activity was seen between fluid from the isthmus and that from the ampulla. Indirect immuno-staining of spermatozoa incubated in the oviductal fluid revealed the association of catalase in the region of the acrosomal cap. Addition of a commercial antibody directed against bovine liver catalase completely inhibited catalase activities from the oviductal fluid. Catalase activity was also detected in porcine oviductal fluid, human oviductal fluid, and cervical mucus. Western blots of oviductal fluid probed with the anti-catalase antibody revealed two major bands at 60 and 40 kDa. An immunoaffinity column was used to purify oviductal catalase, showing a unique band at about 60 kDa when analyzed by SDS-PAGE. The purified protein was incubated with bovine, boar, and human sperm, and Western blots of these sperm after several washes detected a band at 60 kDa, indicating that the protein was bound to sperm membranes. However, bovine liver catalase did not bind to sperm. Since H2O2 is one of the key reactants in the chain reaction of free radical production, this enzyme may play an important role in sperm survival within the female tract.
Lapointe
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Centre de Recherche en Biologie de la Reproduction, Université Laval, Ste-Foy, Québec, Canada.
Sullivan
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Sirard
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MA
D000818:Animals; D001826:Body Fluids / Q000201:enzymology; D002374:Catalase / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D002417:Cattle; D002582:Cervix Mucus / Q000201:enzymology; D004971:Estrus / Q000378:metabolism; D005187:Fallopian Tubes / Q000201:enzymology*; D005260:Female; D006801:Humans; D006861:Hydrogen Peroxide / Q000378:metabolism; D007150:Immunohistochemistry; D066298:In Vitro Techniques; D008297:Male; D008970:Molecular Weight; D017382:Reactive Oxygen Species / Q000378:metabolism; D013084:Sperm-Ovum Interactions / Q000502:physiology; D013094:Spermatozoa / Q000378:metabolism*; D013552:Swine
D017382:Reactive Oxygen Species; D006861:Hydrogen Peroxide; D002374:Catalase