Laboratory of Cell Dynamics, Graduate School of Bioagricultural Sciences, Nagoya University, Nagoya 464-8601, Japan.
J Biochem. 2009 Nov;146(5):617-21. doi: 10.1093/jb/mvp114. Epub 2009 Jul 23.
H(+)-translocating pyrophosphatase converts energy from hydrolysis of pyrophosphate to active H(+) transport across biomembranes. Mutational analysis of Streptomyces coelicolor A3(2) enzyme revealed that amino acid substitution of Phe-388 and Ala-514 altered the enzyme activity. Both residues are located at the interface between the transmembrane domains and cytosolic loops, in which the catalytic domain exists. Systematic amino acid substitution was carried out using the Escherichia coli heterologous expression system. Two of the 38 mutant enzymes, F388Y and A514S, showed a high ratio of H(+)-pump to substrate hydrolysis without decrease in the substrate hydrolysis activity, indicating high energy-coupling efficiency.
H(+)-转运焦磷酸酶将水解焦磷酸盐的能量转化为生物膜上的有效 H(+)转运。对链霉菌 A3(2)酶的突变分析表明，苯丙氨酸 388 和丙氨酸 514 的氨基酸取代改变了酶活性。这两个残基都位于跨膜结构域和胞质环的界面处，催化结构域存在于此。使用大肠杆菌异源表达系统进行了系统的氨基酸取代。38 种突变酶中的两种，F388Y 和 A514S，表现出高的 H(+)-泵与底物水解的比值，而不降低底物水解活性，表明具有高的能量偶联效率。
