Proximal cysteine residue is essential for the enzymatic activities of cytochrome p450cam

Shiro Yoshioka, Satoshi Takahashi, Hiroshi Hori, Koichiro Ishimori, Isao Morishima

研究成果: Article査読

45 被引用数 (Scopus)

抄録

To investigate the functional and structural roles of the proximal thiolate ligand in cytochrome P450cam, we prepared the C357H mutant of the enzyme in which the axial cysteine residue (Cys357) was replaced with a histidine residue. We obtained the unstable C357H mutant by developing a new preparation procedure involving in vitro folding of P450cam from the inclusion bodies. The C357H mutant in the ferrous-CO form exhibited the Soret peak at 420 nm and the Fe-CO stretching line at 498 cm-1, indicating a neutral histidine residue as the axial ligand. However, another internal ligand is coordinated to the heme iron as the sixth ligand in the ferric and ferrous forms of the C357H mutant, suggesting the collapse of the substrate-binding site. The C357H mutant showed no catalytic activity for camphor hydroxylation and the reduced heterolytic/homolytic ratio of the O-O bond scission in the reaction with cumene hydroperoxide. The present observations indicate that the thiolate coordination in P450cam is important for the construction of the heme pocket and the heterolysis of the O-O bond.

本文言語English
ページ(範囲)252-259
ページ数8
ジャーナルEuropean Journal of Biochemistry
268
2
DOI
出版ステータスPublished - 2001
外部発表はい

ASJC Scopus subject areas

  • 生化学

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