title: Assignment of the three disulfide bridges of huwentoxin-I, a neurotoxin from the spider selenocosmia huwena.
abstract: The positions of the disulfide bonds of huwentoxin-I, a neurotoxin from the spider Selenocosmia huwena, have been determined. The existence of three disulfide bonds in the native toxin was demonstrated by mass spectroscopy and the lack of reactivity with a thiol reagent. The assignment procedure involved a combination of tryptic digestion of the native toxin and sequence analysis of both intact and in situ S-carboxymethylated toxin. In situ carboxymethylation is shown to be a useful procedure in sequencing of cysteine- and cystine-containing peptides. Sequence analysis of the intact, cross-linked toxin indicated that no amino acid phenylthiohydantoin (PTH) derivative is seen for the first half-cystine in a cross-linked pair, but that the PTH of dehydroalanine, which can be detected at 313 nm, is seen at the position of the second half-cystine. By sequencing disulfide cross-linked tryptic fragments, the three disulfide linkages in huwentoxin-I could be assigned as Cys2-Cys17, Cys9-Cys22, and Cys16-Cys29.
authors.affiliation: Department of Biology, Hunan Normal University, Changsha, People's Republic of China.
authors.forename: D
authors.initials: D
authors.lastname: Zhang
authors.forename: S
authors.initials: S
authors.lastname: Liang
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D004220:Disulfides / Q000032:analysis*; D010450:Endopeptidases; D013058:Mass Spectrometry; D008969:Molecular Sequence Data; D009498:Neurotoxins / Q000737:chemistry* / Q000302:isolation & purification; D010446:Peptide Fragments / Q000737:chemistry / Q000302:isolation & purification; D011487:Protein Conformation; D030162:Reptilian Proteins; D013111:Spider Venoms / Q000737:chemistry* / Q000302:isolation & purification; D013112:Spiders; D014357:Trypsin
chemical_list: D004220:Disulfides; D009498:Neurotoxins; D010446:Peptide Fragments; D030162:Reptilian Proteins; D013111:Spider Venoms; C083624:huwentoxin I; D010450:Endopeptidases; D014357:Trypsin