Activation of Xenopus eggs by proteases: Possible involvement of a sperm protease in fertilization

Egg activation in cross-fertilization between Xenopus eggs and Cynops sperm may be caused by a protease activity against Boc-Gly-Arg-Arg-MCA in the sp erm acrosome. To determine the role of the sperm protease in fertilization, the protease was purified from Cynops sperm using several chromatographic techniques. We found that purified sperm protease readily hydrolyzes Boc-Gl y-Arg-Arg-MCA and Z-Arg-Arg-MCA, that protease activity was inhibited by th e trypsin inhibitors aprotinin and leupeptin, and that not only the purifie d protease, but also cathepsin B, induces activation in Xenopus eggs. We in seminated unfertilized Xenopus eggs with homologous sperm in the presence o f various peptidyl MCA substrates or protease inhibitors and demonstrated t hat trypsin inhibitors or MCA substrates containing Arg-Arg-RICA reversibly inhibited fertilization of both fully jellied and denuded eggs. Sperm moti lity was not affected by the reagents. An extract obtained from Xenopus spe rm showed hydrolytic activity against Boc-Gly-Arg-Arg-MCA, Z-Arg-Arg-MCA, a nd Arg-MCA. These results suggest that the tryptic protease in Xenopus sper m is involved in fertilization, most likely by participating in egg activat ion, (C) 1999 Academic Press.