FACTOR X-ACTIVATING GLYCOPROTEIN OF RUSSELLS VIPER VENOM - POLYPEPTIDE COMPOSITION AND CHARACTERIZATION OF THE CARBOHYDRATE MOIETIES

There is contradictory information regarding the molecular weight and polypeptide chain composition of RVV-X, a glycoprotein in Russell's vi per venom that is capable of activating factor X to Xa. We show that R VV-X is a 92,880-Da glycoprotein. It consists of three disulfide-linke d polypeptide chains, one heavy chain (alpha-chain, M(r) 57,600) and t wo light chains (beta- and gamma-chains, M(r) 19,400 and 16,400, respe ctively). The two light chains seen on SDS-polyacrylamide gel electrop horesis under reducing conditions are two distinct components of the m olecule, rather than a heterogeneous mixture of a single chain as prev iously reported (Takeya, H., Nishida, S., Miyata, T., Kawada, S., Sais aka, Y., Morita, T., and Iwanaga, S. (1992) J. Biol. Chem. 267, 14109- 14117). The following evidence supports this conclusion. (i) The two l ight chains of RVV-X are present in equal proportion. (ii) The estimat ed molecular weight of an alpha(1) beta(1) gamma(1)-structure closely matches the molecular weight determined by matrix-assisted laser desor ption mass spectrometry. (iii) The amino acid compositions and NH2-ter minal sequences of the beta- and gamma-chains are different. (iv) Alth ough both the beta- and gamma-chains contain one N-linked oligosacchar ide chain each, they are glycosylated differentially. RVV-X contains s ix N-linked oligosaccharides, four in the alpha-chain and one in each of the beta- and gamma-chains. The carbohydrate structures are differe nt from those known for other snake venom glycoproteins, and they rese mble closely those in various mammalian glycoproteins. The majority of the oligosaccharides are complex bi-, tri-, and tetraantennary struct ures, with a small proportion of multiantennary and high mannose-type structures. Two notable structural features of RVV-X oligosaccharides are as follows. (i) Sialic acid residues are linked to beta-galactosyl residues solely by alpha 2,3-linkages, and (ii) bisecting N-acetylglu cosamine residues are present in the majority of the oligosaccharides.