Svedendahl Maria, Hult Karl, Berglund Per
Department of Biochemistry, School of Biotechnology, Royal Institute of Technology (KTH), AlbaNova University Center, SE-106 91 Stockholm, Sweden.
J Am Chem Soc. 2005 Dec 28;127(51):17988-9. doi: 10.1021/ja056660r.
Lipase B from Candida antarctica was redesigned to catalyze the promiscuous reaction of carbon-carbon bond formation. Mutation of the catalytic serine to alanine afforded a mutant that catalyzed Michael additions of 1,3-dicarbonyls to alpha,beta-unsaturated carbonyl compounds at high specific rates, such as 4000 s-1. The enzyme-catalyzed Michael addition reaction followed saturation kinetics and showed substrate inhibition. The designed enzyme showed high rate enhancements with a catalytic proficiency higher than 108, which is on the same level as that observed for enzymes with native substrates.
来自南极假丝酵母的脂肪酶B经过重新设计，以催化碳-碳键形成的混杂反应。将催化丝氨酸突变为丙氨酸得到一个突变体，该突变体能够以高比速率（如4000 s⁻¹）催化1,3-二羰基化合物与α,β-不饱和羰基化合物的迈克尔加成反应。酶催化的迈克尔加成反应遵循饱和动力学并表现出底物抑制。设计的酶显示出高的速率增强，催化效率高于10⁸，这与天然底物酶所观察到的水平相同。
