Elevation of sperm adenosine 3':5'-monophosphate concentrations by a fucose-sulfate-rich complex associated with eggs: I. Structural characterization.
A fucose-sulfate-rich complex (F-SP) capable of causing up to 400-fold elevations of sperm cyclic AMP concentrations was isolated from Strongylocentrotus purpuratus egg jelly and characterized with respect to the relationship of composition to its ability to elevate cyclic AMP. The composition of F-SP varied between different preparations and consisted of, by weight, 32-45% fucose, 36-44% sulfate, 2.5-29% protein, and less than 1% other carbohydrate. The remainder of the weight (approximately 10% in most cases) was accounted for by Na+ or Cs+. The complex caused 45Ca2+ uptake and induced acrosome reactions in addition to its effect on cyclic AMP. The percentage of endogenous protein and sulfate in F-SP were highly correlated with the potency to elevate cyclic AMP with optimal activity observed at the highest relative percentage protein and sulfate. Treatment with NaOH and NaBH4 resulted in the release of most of the protein associated with F-SP but did not reduce sulfate content by more than 1%. The amino acid composition of the total acid hydrolysate did not change after the base treatment, suggesting the absence of serine or threonine O-glycosidic linkages. The NaOH treatment, however, resulted in significant reductions in both the potency and maximal ability of F-SP to elevate cyclic AMP, although it did not totally destroy activity even after treatments for up to 72 h at 37 degrees C. Pronase caused the release of a majority of the protein associated with F-SP and it also significantly reduced both the potency and maximal ability of the complex to elevate cyclic AMP. In contrast to base treatment, HCl (0.1 N) rapidly destroyed F-SP activity and caused a shift in the apparent molecular size of F-SP. Large quantities of fucose were removed from F-SP by the acid prior to a change in the elution position of protein and biological activity was lost, suggesting that the F-SP structure is essential for elevations of cyclic AMP. These results suggest that the fucose-sulfate-rich complex capable of markedly elevating sperm cyclic AMP concentrations requires both protein and sulfate for optimal activity and that protein is either attached to the carbohydrate by a base-labile bond not involving serine or threonine, or is associated with the F-SP structure in a noncovalent manner.
D000596:Amino Acids / Q000032:analysis; D000818:Animals; D002850:Chromatography, Gel; D000242:Cyclic AMP / Q000378:metabolism*; D005260:Female; D005643:Fucose; D008297:Male; D008854:Microscopy, Electron; D010063:Ovum / Q000032:analysis*; D012617:Sea Urchins; D013094:Spermatozoa / Q000378:metabolism*; D013431:Sulfates; D014020:Tissue Extracts / Q000494:pharmacology*
D000596:Amino Acids; D013431:Sulfates; D014020:Tissue Extracts; D005643:Fucose; D000242:Cyclic AMP