硫酶
化学
产量(工程)
克莱森缩合
酿酒酵母
生物催化
丁醇
酶
组合化学
立体化学
有机化学
生物化学
酵母
催化作用
材料科学
反应机理
乙醇
冶金
脱氢酶
作者
Pamela Torres‐Salas,Vicente Bernal,Fernándo López-Gallego,J. Martínez-Crespo,Pedro A. Sánchez‐Murcia,Víctor Hugo Barrera,Rocío Morales-Jiménez,Ana García-Sánchez,Aurora Mañas‐Fernández,Jose Miguel Seoane,Marta Sagrera Polo,Juan de Dios Miranda,Javier Calvo,Sonia Huertas,J. L. Torres,Ana Alcalde-Bascones,Sergio González‐Barrera,Federico Gago,Antonio Morreale,María del Mar González-Barroso
出处
期刊:Biochemistry
[American Chemical Society]
日期:2018-02-06
卷期号:57 (8): 1338-1348
被引量:9
标识
DOI:10.1021/acs.biochem.7b01186
摘要
Thiolases catalyze the condensation of acyl-CoA thioesters through the Claisen condensation reaction. The best described enzymes usually yield linear condensation products. Using a combined computational/experimental approach, and guided by structural information, we have studied the potential of thiolases to synthesize branched compounds. We have identified a bulky residue located at the active site that blocks proper accommodation of substrates longer than acetyl-CoA. Amino acid replacements at such a position exert effects on the activity and product selectivity of the enzymes that are highly dependent on a protein scaffold. Among the set of five thiolases studied, Erg10 thiolase from Saccharomyces cerevisiae showed no acetyl-CoA/butyryl-CoA branched condensation activity, but variants at position F293 resulted the most active and selective biocatalysts for this reaction. This is the first time that a thiolase has been engineered to synthesize branched compounds. These novel enzymes enrich the toolbox of combinatorial (bio)chemistry, paving the way for manufacturing a variety of α-substituted synthons. As a proof of concept, we have engineered Clostridium’s 1-butanol pathway to obtain 2-ethyl-1-butanol, an alcohol that is interesting as a branched model compound.
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