On the macromolecular composition of the zona pellucida from porcine oocytes.
The chemical and immunological relation between the glycoprotein components of the pig oocyte zona pellucida resolved by two-dimensional polyacrylamide gel electrophoresis was investigated. After disulfide bond reduction, four microheterogeneous glycoprotein components with apparent molecular weights of 25K, 55K, 65K, and 90K were resolved. When disulfide bonds were left unreduced, two microheterogeneous glycoprotein components were resolved with apparent molecular weights of 55K and 90K. Actin was present, but as a contaminant of the zona pellucida rather than as a true component. The structural relation of these components was investigated using deglycosylation with trifluoromethane-sulfonic acid, limited proteolysis with Staphylococcus aureus V8 protease, amino acid and carbohydrate composition analyses, sequence analysis, and monoclonal antibodies. The 25K and 65K components comigrated with the 90K component when disulfide bonds were not reduced. When the intermolecular disulfide bonds crosslinking the two components were reduced, the 25K and 65K components behaved independently. The 25K and 65K components were derived from the 90K glycoprotein family by proteolysis. The 25K component originated from the C-terminal end, and the 65K component from the N-terminal end of the 90K glycoprotein. The 55K component was composed of two chemically and antigenically distinct glycoproteins, termed 55K alpha and 55K beta, that electrophoretically comigrated. The N-terminal amino acid of the 55K alpha family was blocked. The 55K beta family had an N-terminal amino acid sequence of Asp-Val-Pro-Thr-Ile-Gly-Leu-Ser-X-Ala-Pro-Thr. Thus, the two to four electrophoretic components of the zona pellucida observed on gel electrophoresis are derived from three glycoprotein families.
D000199:Actins / Q000032:analysis; D000595:Amino Acid Sequence; D000818:Animals; D004220:Disulfides; D004591:Electrophoresis, Polyacrylamide Gel; D010450:Endopeptidases; D005260:Female; D006023:Glycoproteins / Q000032:analysis*; D006031:Glycosylation; D046911:Macromolecular Substances; D008970:Molecular Weight; D009865:Oocytes / Q000648:ultrastructure*; D010063:Ovum / Q000032:analysis*; D010084:Oxidation-Reduction; D010446:Peptide Fragments / Q000032:analysis; D012697:Serine Endopeptidases; D013552:Swine; D015044:Zona Pellucida / Q000032:analysis*
D000199:Actins; D004220:Disulfides; D006023:Glycoproteins; D046911:Macromolecular Substances; D010446:Peptide Fragments; D010450:Endopeptidases; D012697:Serine Endopeptidases; C028344:glutamyl endopeptidase