Role of Carboxyl Residues Surrounding Heme of Human Cytochrome b₅in the Electrostatic Interaction with NADH-Cytochrome b₅Reductase

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• 作者：Kawano ; Masaya ; Shirabe ; Komei ; Nagai ; Takushi ; Takeshita ; Masazumi
• 刊名：Biochemical and Biophysical Research Communications
• 出版年：1998
• 出版时间：April 28, 1998
• 年：1998
• 卷：245
• 期：3
• 页码：666-669
• 全文大小：97.0 K

文摘

To identify the cytochrome b₅residues responsible for the electrostatic interaction with NADH-cytochrome b₅reductase (b₅R), we prepared and characterized the cytochrome b₅mutants in which Glu41, Glu42, Glu63, Asp70, and Glu73 were replaced by Ala, utilizing site-directed mutagenesis and the expression system for cytochrome b₅inEscherichia coli.Apparent Km values of the wild type b₅R for Glu42Ala cytochrome b₅and Asp70Ala cytochrome b₅were approximately three-fold and six-fold higher than that for the wild type cytochrome b₅, respectively, while the kcat values for those mutants were not remarkably affected. In contrast, Glu41Ala, Glu63Ala, and Glu73Ala cytochrome b₅showed almost the same kinetic properties as the wild type cytochrome b₅. Furthermore, kinetic studies on combinations of the cytochrome b₅and b₅R mutants suggested the interaction between Glu42 and Asp70 of cytochrome b₅and Lys125 and Lys41 of b₅R, respectively, in the reaction.