Release of ovoperoxidase from sea urchin eggs hardens the fertilization membrane with tyrosine crosslinks.
One feature of fertilization is the alteration of the vitelline layer, by components released from the egg, to produce an elevated, covalently crosslinked, hard, insoluble, fertilization membrane. The following evidence indicates that crosslinking and hardening are caused by the production of diand trityrosyl residues, by oxidation of protein-bound tyrosyl residues in the presence of a peroxidase. Hardening of the fertilization membrane, as evidenced by its loss of solubility in 50 mM dithiothreitol, is inhibited by compounds known to inhibit many peroxidases. A peroxidase, here called the ovoperoxidase, is released from eggs at fertilization. This enzyme is inhibited by the same compounds that inhibit hardening and at similar concentrations. Inhibitors of the ovoperoxidase and the hardening reaction include KCN, 3-amino-1,2,4-triazole, NaN(3), phenylhydrazine, K(4)Fe(CN)(6), sodium sulfite, and glycine ethyl ester. In addition, tyramine and N-acetyltyrosine both inhibit hardening, but O-methyltyrosine does not. Dityrosyl and trityrosyl residues are found in acid hydrolysates of isolated, hardened fertilization membranes. These residues have been identified by cellulose phosphate column chromatography, thin-layer chromatography, and amino acid analysis. The amino acid data have been used to estimate that there is one dityrosine crosslink per 55,000 daltons of protein. We suggest that, by catalyzing the crosslinking of tyrosyl residues, the ovoperoxidase leads to the production of a hard fertilization membrane that blocks the entry of additional sperm. Because peroxidases are spermicidal, a secondary function of the enzyme could be to kill sperm in the vicinity of the fertilized egg.
D000596:Amino Acids / Q000032:analysis; D000818:Animals; D005260:Female; D005306:Fertilization; D006244:Hardness; D010063:Ovum / Q000201:enzymology*; D010544:Peroxidases / Q000378:metabolism*; D012617:Sea Urchins; D014443:Tyrosine / Q000032:analysis / Q000502:physiology*; D014817:Vitelline Membrane / Q000378:metabolism* / Q000502:physiology / Q000648:ultrastructure
D000596:Amino Acids; D014443:Tyrosine; D010544:Peroxidases
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