Effect of site‐directed mutagenesis on conserved positions of Drosophila alcohol dehydrogenase

FEBS Letters

Atrian, Sílvia¹  Cols, Neus¹  Gonzàlez-Duarte, Roser¹  Marfany, Gemma¹ 

[1] Department de Genètica, Facultat de Biologia, Universität de Barcelona, Av. Diagonal 645, E-08028 Barcelona, Spain

关键词: Site-directed mutagenesis;    Active site residue;    Alcohol dehydrogenase;    Short-chain dehydrogenase;    Drosophila;   

DOI  :  10.1016/0014-5793(93)80043-T

学科分类：生物化学/生物物理

来源: John Wiley & Sons Ltd.

【 摘 要 】

Tyr¹⁵² and Lys¹⁵⁶ may be functionally important residues in Drosophila ADH as they are conserved in the genus and in all short-chain dehydrogenases. In addition, unaltered Gly positions could have a crucial role in the building of the structural framework. We have modified Drosophila ADH and expressed the mutant forms in E. coli. Mutation of Tyr¹⁵² to Glu or Gin, Lys¹⁵⁶ to Ile, Gly¹⁸⁴ to Leu, and the double mutant Gly¹³⁰ to Cys and Gly¹³³ to Ile, all rendered, with different substrates and at different pHs, an inactive enzyme. Results suggest that Tyr¹⁵² and Lys¹⁵⁶are involved in catalysis and that Gly¹³⁰, Gly¹³³ and Gly¹⁸⁴ contribute substantially to the structure of the active form.

【 授权许可】

Unknown