ASSIGNMENT OF DISULFIDE BOND LOCATION IN PROTHORACICOTROPIC HORMONE OF THE SILKWORM, BOMBYX-MORI - A HOMODIMERIC PEPTIDE

被引:56
作者
ISHIBASHI, J
KATAOKA, H
ISOGAI, A
KAWAKAMI, A
SAEGUSA, H
YAGI, Y
MIZOGUCHI, A
ISHIZAKI, H
SUZUKI, A
机构
[1] UNIV TOKYO, FAC AGR, DEPT AGR CHEM, BUNKYO KU, TOKYO 113, JAPAN
[2] NAGOYA UNIV, SCH SCI, DEPT BIOL, NAGOYA 46401, JAPAN
关键词
D O I
10.1021/bi00185a031
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
The disulfide bond location of a homodimeric peptide, prothoracicotropic hormone (PTTH) of the silkworm, Bombyx mori, was determined by a combination of partial reduction and sequence analysis of peptide fragments generated through a partial reduction of PTTH followed by alkylation and enzyme digestion. The partial reduction and S-alkylation broke the interchain disulfide bond but did not affect the intrachain disulfide bonds, generating monomeric PTTH whose intrachain disulfide bonds were kept intact. This monomeric PTTH has about one-half the biological activity of intact PTTH. Sequence analysis of the fragments generated by lysyl endopeptidase digestion of this monomeric PTTH after complete reduction and S-alkylation by another S-alkylating reagent showed that only the Cys15 residue was reduced and S-alkylated by the foregoing partial reduction, indicating that this residue formed the interchain disulfide bond. The other disulfide bonds which formed intrachain bridgings were determined by sequence and mass analyses of the fragments generated by two successive enzyme digestions of the monomeric PTTH. In conclusion, the disulfide bond location of PTTH was assigned to Cys15-Cys15' as an interchain disulfide linkage and Cys17-Cys54, Cys40-Cys96, and Cys48-Cys98 as intrachain disulfide linkages.
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页码:5912 / 5919
页数:8
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