Species-specific aggregation factor in sponges. Sialyltransferase associated with aggregation factor.
The sialyltransferase (= glycoprotein-sialic acid transferase) was studied in the sponge Geodia cydonium, a mesozoan organism. The experiments were performed both in intact cellular and in isolated enzyme systems. It is shown, that desialylated cells show a lower aggregation potency than the controls. During aggregation enzymic sialylation of desialylated sponge cells occurs in the presence of an aggregation factor, which is associated with a high molecular weight particle. The sialylation process is temperature-dependent and can be inhibited by N-ethylmaleimide. Sialylation occurs predominantly at a distinct cell surface component, the aggregation receptor. The sialyltransferase was isolated and purified by the following steps: Sepharose 4B, CM-cellulose, Nonidet treatment, and Sephadex G-100. By this procedure the enzyme was purified 680-fold with a 31% yield. The sialyltransferase is originally associated with the high molecular weight particle also carrying the aggregation factor. In the last step the aggregation factor was separated from the sialyltransferase. The enzyme catalyzes the transfer of sialic acid from CMP-sialic acid to the desialylated aggregation receptor. The molecular weight of the sialyltransferase has been determined to be 52,000. Kinetic studies revealed no lag phase and a dependence on enzyme concentration. The purified transferase has a pH optimum of 7.75 and requires 200 mM NaCl for activity. No requirement for Mg2+ or Ca2+ could be observed. The reaction is inhibited by 10 micronM N-ethylmaleimide.
D000818:Animals; D002118:Calcium / Q000494:pharmacology; D015815:Cell Adhesion Molecules; D002449:Cell Aggregation / Q000187:drug effects*; D005033:Ethylmaleimide / Q000494:pharmacology; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008274:Magnesium / Q000494:pharmacology; D008970:Molecular Weight; D009439:Neuraminidase / Q000494:pharmacology; D011161:Porifera / Q000201:enzymology*; D011506:Proteins / Q000302:isolation & purification / Q000502:physiology; D011955:Receptors, Drug; D012794:Sialic Acids / Q000378:metabolism; D012799:Sialyltransferases / Q000302:isolation & purification / Q000378:metabolism*; D012965:Sodium Chloride / Q000494:pharmacology; D013045:Species Specificity; D014166:Transferases / Q000378:metabolism*
D015815:Cell Adhesion Molecules; D011506:Proteins; D011955:Receptors, Drug; D012794:Sialic Acids; C013065:cell aggregation factors; D012965:Sodium Chloride; D014166:Transferases; D012799:Sialyltransferases; D009439:Neuraminidase; D008274:Magnesium; D005033:Ethylmaleimide; D002118:Calcium