Isolation, characterization, and immunochemical properties of a giant protein from sea urchin egg cytomatrix

Exp Cell Res. 1990 Aug;189(2):253-60. doi: 10.1016/0014-4827(90)90244-5.

Abstract

A giant protein of apparent molecular weight (Mr) 2000 kDa, as determined by SDS-PAGE, was isolated and partially purified, under denaturing conditions, from the detergent-resistant cytomatrix of unfertilized sea urchin egg. Immunoblot analysis and indirect immunofluorescence microscopy observations indicated that this high-molecular-weight protein cross-reacted with the immunospecific serum raised against chicken breast muscle beta-connectin. However, rotary-shadowing electron microscopy images of the protein revealed short threadlike structures which appear morphologically different from beta-connectin structure. Indirect immunofluorescence localization of the protein with anti-beta-connectin serum showed a distribution throughout the whole unfertilized egg cytomatrix. This immunofluorescence pattern seems to change upon egg fertilization, since at metaphase the fluorescence stain appears to be excluded from the mitotic apparatus region as revealed by the double immunolabeling with anti-beta-connectin serum and monoclonal anti-alpha-tubulin antibody. Moreover, when egg cortical fragments were double-labeled with anti-beta-connectin serum and rhodamin-conjugated phalloidin, it was observed that the microfilaments assembled after fertilization seem to be in close association with the protein at the cleavage furrow and other locations. The possible significance of this sea urchin egg connectin(titin)-like protein is discussed.

MeSH terms

  • Animals
  • Electrophoresis, Polyacrylamide Gel
  • Female
  • Male
  • Microscopy, Electron
  • Microscopy, Fluorescence
  • Molecular Weight
  • Ovum / analysis*
  • Ovum / cytology
  • Proteins / analysis
  • Proteins / isolation & purification*
  • Proteins / ultrastructure
  • Sea Urchins
  • Ultracentrifugation
  • Zygote / analysis
  • Zygote / cytology

Substances

  • Proteins