Structural studies of a major hemorrhagin (rhodostoxin) from the venom of Calloselasma rhodostoma (Malayan pit viper).
The complete amino acid sequence, disulfide linkages, glycosylation sites, and carbohydrate structure of rhodostoxin, the major hemorrhagin from Calloselasma rhodostoma (Malayan pit viper), have been determined. This sequence confirmed the deduced amino acid sequence of the putative hemorrhagic protein encoded by the prorhodostomin cDNA of C. Rhodostoma. Rhodostoxin contained four disulfide bonds that link Cys19-Cys60, Cys117-Cys198, Cys157-Cys182, and Cys159-Cys165. It is the first four-disulfide proteinase reported among all known venom metalloproteinases, which are either of the two-disulfide or three-disulfide type. Peptide-mapping and dot-blotting experiments showed the presence of two glycopeptides. Subsequent sequencing of these peptides established that the N-glycosylation sites are located at residues 91 and 181 of the amino acid sequence of the matured protein. Mass spectrometric analyses of these glycopeptides showed that they contain an oligosaccharide structure consisting of 4 units of N-acetylglucosamine, 5 units of hexose, 1 unit of fucose, and 2 units of neuraminic acids. The complete carbohydrate structure was then established by 2-D mapping analysis of the pyridylamino-oligosaccharides after hydrazinolysis and pyridylamination of the glycan chains.
Chung
M C
MC
Department of Biochemistry and Bioprocessing Technology Centre, National University of Singapore, Republic of Singapore.
Ponnudurai
G
G
Kataoka
M
M
Shimizu
S
S
Tan
N H
NH
D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D002240:Carbohydrate Sequence; D002851:Chromatography, High Pressure Liquid; D003435:Crotalid Venoms / Q000737:chemistry* / Q000235:genetics; D004220:Disulfides / Q000737:chemistry; D010450:Endopeptidases / Q000737:chemistry* / Q000235:genetics; D006023:Glycoproteins / Q000737:chemistry* / Q000235:genetics; D006031:Glycosylation; D013058:Mass Spectrometry; D008969:Molecular Sequence Data; D015394:Molecular Structure; D009844:Oligosaccharides / Q000737:chemistry / Q000235:genetics; D010802:Phylogeny; D011134:Polysaccharides / Q000737:chemistry / Q000235:genetics
D003435:Crotalid Venoms; D004220:Disulfides; D006023:Glycoproteins; D009844:Oligosaccharides; D011134:Polysaccharides; C097743:rhodostoxin protein, Calloselasma rhodostoma; D010450:Endopeptidases; C041605:hemorrhagic proteinase IV