Analysis of 46-kDa protein in the perivitelline membrane of Japanese quail (Coturnix japonica)

The extracellular matrix surrounding the oocyte before ovulation is called the perivitelline membrane (PL) in avian species. The PL is constructed with two major glycoproteins, ZPC and ZP1, which are synthesized in the ovarian granulosa cells and the liver, respectively. Although the properties of the major components in the PL have been examined, knowledge about the nature of its minor constituents is lacking. In this study we focused on PL protein, which migrates at 46-kDa in the gel of SDS-PAGE. N-terminal sequence analysis demonstrated that the 46-kDa protein is the C-terminal fragment of ZP1. Analysis of lysylendopeptidase digests or cyanogens bromide-degraded fragments of ZP1 confirmed this postulate. Western blot analysis using antiserum against 46-kDa protein indicated the absence of 46-kDa protein in the serum. Moreover, small immunoreactive bands, thought to be cleaved fragments of ZP1, were detected in the PL lysate by western blot analysis using antiserum against the N-terminal peptide of ZP1. These results indicated that the N-terminal proteolytic processing of ZP1 might take place after the arrival of ZP1 at the ovary, and the resulting product, 46-kDa protein, is incorporated into the PL.