BINDING OF A BOVINE OVIDUCTAL FLUID CATALASE TO MAMMALIAN SPERMATOZOA

Hydrogen peroxide (H2O2) is a reactive oxygen species that at low conc entration is toxic to sperm. H2O2 inhibits not only sperm viability bu t also the acrosome reaction, sperm-egg binding, and oocyte penetratio n. 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 survi val, we hypothesized that it might secrete catalase. We found that ovi ductal fluid, harvested from washed cells collected at the slaughterho use, possessed catalase-specific activity that varied during the estro us cycle. Catalase activity increased during the cycle and reached its maximal level just before ovulation (Days 18-20). No significant diff erence in activity was seen between fluid from the isthmus and that fr om the ampulla. Indirect immunostaining of spermatozoa incubated in th e oviductal fluid revealed the association of catalase in the region o f the acrosomal cap. Addition of a commercial antibody directed agains t bovine liver catalase completely inhibited catalase activities from the oviductal fluid. Catalase activity was also detected in porcine ov iductal fluid, human oviductal fluid, and cervical mucus. Western blot s of oviductal fluid probed with the anti-catalase antibody revealed t wo 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 boun d to sperm membranes. However, bovine liver catalase did not bind to s perm. Since H2O2 is one of the key reactants in the chain reaction of free radical production, this enzyme may play an important role in spe rm survival within the female tract.