Fig. 1. Tyrosine phosphorylation of Xenopus egg proteins in response to fertilization, Ca2+ ionophore and H2O2. SDS-solubilized extract was prepared from Xenopus eggs that had been treated with either sperm (insemination, lanes 1-6), A23187 (0.5 µM, lanes 7-12) or H2O2 (10 mM, lanes 13-18) for the indicated times. Egg proteins (each lane contains 50 µg protein that corresponds to one egg) were separated by SDS-PAGE and analyzed by immunoblotting (IB) with either (A) anti-phosphotyrosine antibody (PY99, 1 µg/ml), (B) anti-phosphotyrosine antibody plus 10 mM L-phosphotyrosine or (C) anti-pY416 antibody (10 µg/ml). Asterisks in A indicate the positions of tyrosine-phosphorylated proteins. Black and white arrowheads in A indicate the positions of a 90 kDa protein and a 42 kDa MAP kinase, respectively. Black arrowheads in C indicate the position of Xyk (57 kDa and partially degraded 55 kDa bands). Prestained molecular size markers are maltose binding protein (MBP)-fusion ß-galactosidase (175 kDa), MBP-fusion paramyosin (80 kDa), glutamic dehydrogenase (62 kDa), aldolase (47.5 kDa) and triosephosphate isomerase (32.5 kDa).
