Core sugar residues of the N-linked oligosaccharides of Russell's viper venom factor X-activator maintain functionally active polypeptide structure.
We have previously showed that factor X activator of Russell's viper venom (RVV-X) contains six N-linked oligosaccharide chains: four in the heavy chain and one in each of the two light chains [Gowda, D.C., Jackson, C.M., Hensley, P., & Davidson, E.A. (1994) J. Biol. Chem. 269, 10644-10650]. In the present study, we have investigated the role of the carbohydrate moieties in the structure and functional activity of RVV-X. Sequential removal of sugar residues from the terminal ends by exoglycosidases, up to 50% of total carbohydrates, did not significantly alter the activity of RVV-X, demonstrating that the peripheral carbohydrate moieties are not involved in interactions with factor X. However, removal of whole oligosaccharide chains by N-glycanase caused an almost total loss of the ability of RVV-X to activate factor X to factor Xa. In parallel with these observations, circular dichroism spectroscopy showed that complete deglycosylation, but not the removal of peripheral sugars, caused a significant change in the secondary structure. Together, these data demonstrate that the oligosaccharide chains are necessary for the functional activity, and that the trimannosylchitobiose core residues are sufficient for the maintenance of the native polypeptide structure.
Gowda
D C
DC
Department of Biochemistry and Molecular Biology, Georgetown University Medical Center, Washington, DC 20007, USA.
Jackson
C M
CM
Kurzban
G P
GP
McPhie
P
P
Davidson
E A
EA
D000818:Animals; D002241:Carbohydrates / Q000737:chemistry; D002417:Cattle; D002942:Circular Dichroism; D005170:Factor X / Q000378:metabolism; D006026:Glycoside Hydrolases; D006031:Glycosylation; D066298:In Vitro Techniques; D008666:Metalloendopeptidases / Q000737:chemistry* / Q000378:metabolism; D015394:Molecular Structure; D009844:Oligosaccharides / Q000737:chemistry; D017433:Protein Structure, Secondary
D002241:Carbohydrates; D009844:Oligosaccharides; D005170:Factor X; D006026:Glycoside Hydrolases; D008666:Metalloendopeptidases; C033858:russellysin