Identification of avidin‐interacting proteins in Spisula oocytes: Correlation with germinal vesicle breakdown

Abstract

AbstractPeroxidase‐conjugated avidin was used to determine the presence of avidin‐interacting proteins in Spisula oocytes. By a transblot method, avidin‐peroxidase conjugate interacted with the two proteins with estimated molecular weights of 70,000 and 118,000 in the cytosolic fraction of Spisula oocyte. The reaction with the same proteins in the particulate fraction was considerably reduced. None or weak reaction occurred with proteins prepared from Spisula muscle, sperm, and body fluid. With Arbacia oocytes or hen egg‐yolk and egg‐white proteins, the reaction was minimal or none, respectively. When the avidin‐peroxidase conjugate was preincubated with biotin, its stain‐ ing activity was lost, suggesting that the interacting proteins probably contain biotin.To determine whether avidin‐interacting proteins are involved in oocyte maturation, Spisula oocytes were treated with 50 mM KCl or 5 μM serotonin to induce germinal vesicle breakdown (GVBD). The staining reaction with the 70,000‐ and 118,000‐dalton proteins in the samples prepared from oocytes treated with KCl or serotonin was greatly intensified and an additional protein with an estimated molecular weight of 220,000 was stained. The occurrence of avidin‐interacting proteins in Spisula oocytes was further verified by the positive staining reaction with a cytochemical method using fluorescein isothiocyanate (FITC)‐conjugated avidin.