Escherichia coli inorganic pyrophosphatase : site‐directed mutagenesis of the metal binding sites

FEBS Letters

Rodina, Elena²  Harutyunyan, Emil¹  Oganessyan, Vaheh¹  Nazarova, Tatjana²  Kurilova, Svetlana²  Avaeva, Svetlana²  Vorobyeva, Natalja³  Ignatov, Pavel³ 

[1] Institute of Crystallography, Russian Academy of Science, Moscow, Russian Federation;A.N Belozersky Institute of Physico-Chemical Biology, Moscow State University, Moscow 119899, Russian Federation;Department of Chemistry, Moscow State University, Moscow, Russian Federation

关键词: Inorganic pyrophosphatase;    Aspartic acid;    Site-directed mutagenesis;    Kinetics;    Differential spectrophotometry;   

DOI  :  10.1016/S0014-5793(96)01296-3

学科分类：生物化学/生物物理

来源: John Wiley & Sons Ltd.

【 摘 要 】

Aspartic acids 65, 67, 70, 97 and 102 in the inorganic pyrophosphatase of Escherichia coli, identified as evolutionarily conserved residues of the active site, have been replaced by asparagine. Each mutation was found to decrease the κ ^app value by approx. 2–3 orders of magnitude. At the same time, the K _m values changed only slightly. Only minor changes take place in the pK values of the residues essential for both substrate binding and catalysis. All mutant variants have practically the same affinity to Mg²⁺ as the wild-type pyrophosphatase.

【 授权许可】

Unknown