Analysis of the reaction mechanism of the non‐specific endonuclease of Serratia marcescens using an artificial minimal substrate

FEBS Letters

Franke, Ingo¹  Friedhoff, Peter¹  Kolmes, Bettina¹  Pingoud, Alfred¹ 

[1]Institut für Biochemie, Fachbereich Biologie, Justus-Liebig-Universität, Heinrich-Buff-Ring 58, D-35392 Giessen, Germany

关键词: DNase 1;    Staphylococcal nuclease;    Phosphodiesterase;    Enzyme mechanism;    Acid base catalysis;    Deoxythymidine 3′;    5′-bis-(p-nitrophenyl- phosphate);   

DOI  :  10.1016/S0014-5793(96)01210-0

学科分类：生物化学/生物物理

来源: John Wiley & Sons Ltd.

【 摘 要 】

We have studied the mechanism of action of the Serratia nuclease using deoxythymidine 3′,5′-bis-(p-nitrophenyl-phosphate) as a substrate. A comparison of the activity with which the wild-type enzyme and several mutant enzymes attack this artificial substrate and herring sperm DNA, respectively, supports the suggestion that His⁸⁹ is the general base and a Mg²⁺ ion bound to Glu¹²⁷ the general acid in the mechanism of phosphodiester bond hydrolysis by the Serratia nuclease, and that Asn¹¹⁹ directly participates in catalysis, for example by transition state stabilisation. Arg⁵⁷, Arg⁸⁷ and Arg¹³¹, essential for nuclease activity, are not needed for cleavage of the artificial substrate, suggesting that they are involved in binding and positioning of nucleic acid substrates.

【 授权许可】

Unknown