Assignment of the three disulfide bonds of Selenocosmia huwena lectin-I from the venom of spider Selenocosmia huwena.
The positions of the disulfide bonds of Selenocosmia huwena lectin-I (SHL-I) from the venom of the Chinese bird spider S. huwena have been determined. The existence of three disulfide bonds in the native SHL-I was proved by matrix-assisted laser desorption ionization time-of-flight mass spectroscopic analysis. To map the disulfide bonds, native SHL-I was proteolytically digested. The resulting peptides were separated by reverse phase high-performance liquid chromatography. Matrix-assisted laser desorption ionization time-of-flight mass spectroscopic analysis indicated the presence of one disulfide bond Cys7-Cys19. The partially reduced peptides by using Tris-(2-carboxyethyl)-phosphine at pH 3.0 were purified by reverse phase high-performance liquid chromatography. Four M Guanidine-HCl was found to increase the yields of partially reduced peptides prominently. The free thiols were carboxamidomethlate by iodoacetamide. The specific location of another disulfide bond Cys2-Cys14 was proved by comparing N-terminal sequencing analysis of the partially reduced and alkylated SHL-I with that of the intact peptide. Finally, the three disulfide linkage of SHL-I could be assigned as Cys2-Cys14, Cys7-Cys19, Cys13-Cys26.
Li
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Department of Biology, Human Normal University, Changsha, People's Republic of China.
Liang
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D000595:Amino Acid Sequence; D002851:Chromatography, High Pressure Liquid; D002852:Chromatography, Ion Exchange; D004220:Disulfides / Q000737:chemistry*; D019476:Insect Proteins; D037102:Lectins / Q000737:chemistry* / Q000302:isolation & purification; D008969:Molecular Sequence Data; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D013111:Spider Venoms / Q000737:chemistry*
D004220:Disulfides; D019476:Insect Proteins; D037102:Lectins; D013111:Spider Venoms; C098229:lectin-like peptide-I, Selenocosmia huwena