Binding sites of -SH reagents in dividing sea urchin egg

Abstract

Inhibition of cleavage of sea urchin eggs by -SH reagents was examined by varying their concentration, and the time and stage of the treatment during the first cleavage cycle. It was found that more than half of -SH groups in the ‘cortex protein’, localized in the cortical layer, reacted with p-chloromercuribenzoate (PCMB) when cleavage was suppressed, while other protein fractions of eggs revealed only a little binding of PCMB. The amount of non-protein -SH groups (NPSH) in the egg decreased to a level 50% that of the intact cell when cytokinesis was completely blocked by 1 mM PCMB. Even at lower PCMB conc., which did not block cell division, the NPSH level decreased to 60% and remained there. In pulse-treatment with 0.1 mM N-ethylmaleimide (NEM), which did not block cell division, NPSH groups reacted quickly with the reagent to protect protein -SH groups from the alkylation. However, if the concentration was increased, the pulse-treatment resulted in suppression of cleavage and alkylation of protein -SH groups, especially of cortex protein, accompanied by decrease in ATPase activity involved in the cortex protein fraction.