Enhancing the atypical esterase promiscuity of the γ-lactamase Sspg from Sulfolobus solfataricus by substrate screening
Enhancing the atypical esterase promiscuity of the γ-lactamase Sspg from Sulfolobus solfataricus by substrate screening
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DOI：
10.1007/s00253-019-09758-3
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发表时间：
2019
期刊：
影响因子：
--
通讯作者：
Sheng Wu
中科院分区：
文献类型：
--
作者：
Jianjun Wang;Hongtao Zhao;Guogang Zhao;Dunfu Chen;Yong Tao;Sheng Wu
Promiscuous enzymes can be modified by protein engineering, which enables the catalysis of non-native substrates. γ-lactamase.Sspg from Sulfolobus solfataricus is an enzyme with high activity, high stability, and pronounced tolerance of high concentrations of the γ-lactam substrate. These characteristics suggest Sspg as a robust enzymatic catalyst for the preparation of optically.pure γ-lactam. This study investigated the modification of this enzyme to expand its application toward resolving chiral esters. γLactamase-esterase conversion was performed by employing a three-step method: initial sequence alignment, followed by.substrate screening, and protein engineering based on the obtained substrate-enzyme docking results. This process of finetuning of chemical groups on substrates has been termed Bsubstrate screening.^ Steric hindrance and chemical reactivity of.the substrate are major concerns during this step, since both are determining factors for the enzyme-substrate interaction. By.employing this three-step method, γ-lactamase Sspg was successfully converted into an esterase with high enantioselectivity.towards phenylglycidate substrates (E value > 300). However, since both wild-type Sspg and Sspg mutants did not hydrolyze.para-nitrophenyl substrates (pNPs), this esterase activity was termed Batypical esterase activity.^ The γ-lactamase activity and.stability of the Sspg mutants were not severely compromised. The proposed method can be applied to find novel multi-functional.enzyme catalysts within existing enzyme pools.
