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One feature of fertilization is the alteration of the vitelline layer, by components released from the e, 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 thepresence 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, NaN3, phenylhydrazine, K4Fe(CN)o, 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. Fertilization involves the union of two haploid genomes to make a new diploid individual. To ensure that only one sperm fertilizes an egg, many eggs become modified after penetration by the first sperm. In the case of the sea urchin Strongylocentrotus purpuratus, an immediate block to further sperm entry (polyspermy) is caused by a depolarization of the plasma membrane of the egg (1). After depolarization, the egg surface is rearranged by the cortical reaction (reviewed in refs. 2 and 3), in which secretory vesicles beneath the plasma membrane of the unfertilized egg release their contents in a massive exocytosis. A trypsin-like protease is released to cleave sperm receptors from the egg and thus serve as a second block. to polyspermy (3-5). During the cortical reaction, the vitelline layer that surrounds the egg elevates and is converted into a fertilization membrane. The fertilization membrane becomes altered in its morphology, solubility, and permeability in a discrete, multistep, assembly process that occurs in fixed sequence (6). This elevated structure is another block to sperm entry.The fertilization membrane is formed from the vitelline layer in a reaction with components released from the cortical granules (2, 3, 6). The structure thus produced is resistant to physical deformation (7), solubilization by protein denaturants (...

Some children and adolescents with sickle cell disease experience frequent painful episodes. To gain information about the natural history of the pain and its impact on sleep and school attendance, we developed a home-based diary system. Eighteen children and adolescents completed 4756 diary days, with an average compliance of 75%. Pain was reported on 30% of days and was managed at home nine-tenths of the time. Girls reported more days with pain than did boys, and age was positively correlated with the length of the painful episodes. The pain affected school attendance and sleep. Patients were absent from school on 21% of 3186 school days, with half of the absenteeisms on days with reported pain. Of the pain-associated absenteeisms, two-thirds occurred when pain was managed at home, and one-third when patients were hospitalized. The average consecutive number of school days missed was 2.7. These findings have implications for developmentally critical activities.