Histidine 64 is not required for high CO2 hydration activity of human carbonic anhydrase II

FEBS Letters

Behravan, Gity¹  Wallgren, Katarina¹  Sandström, Jan¹  Liang, Zhi-wei¹  Forsman, Cecilia¹  Jonsson, Bengt-Harald¹  Lindskog, Sven¹  Ren, Xilin¹ 

[1] Department of Biochemistry, University of Umeå, S-901 87 Umeå, Sweden

关键词: Carbonic anhydrase;    Site-directed mutagenesis;    Proton transfer;    Catalytic mechanism;   

DOI  :  10.1016/0014-5793(88)81156-6

学科分类：生物化学/生物物理

来源: John Wiley & Sons Ltd.

【 摘 要 】

To test the hypothesis that histidine 64 in carbonic anhydrase II has a crucial role as a ‘proton shuttle group’ during catalysis of CO₂-HCO⁻ ₃ interconversion, this residue was replaced by lysine, glutamine, glutamic acid and alanine by site-directed mutagenesis. All these variants turned out to have high CO₂ hydration activities. The k _cat values at pH 8.8 and 25°C were only reduced by 1.5–3.5-fold compared to the unmodified enzyme. These results show that intramolecular proton transfer via His 64 is not a dominating pathway in the catalytic reaction. The variants also catalyze the hydrolysis of 4-nitrophenyl acetate. The pK _a values for the activity-controlling group are between 6.8 and 7.0 for all studied forms of the enzyme except the Glu 64 variant which shows a complex pH dependence with the major pK _a shifted to 8.4

【 授权许可】

Unknown