一组钴和铁依赖型腈水合酶的功能表达和特性研究。
Functional Expression and Characterization of a Panel of Cobalt and Iron-Dependent Nitrile Hydratases.
机构信息
Austrian Center of Industrial Biotechnology GmbH, 8010 Graz, Austria.
Molecular Science Institute, School of Chemistry, University of the Witwatersrand, Johannesburg P.O. Wits 2050, South Africa.
出版信息
Molecules. 2020 May 28;25(11):2521. doi: 10.3390/molecules25112521.
Nitrile hydratases (NHase) catalyze the hydration of nitriles to the corresponding amides. We report on the heterologous expression of various nitrile hydratases. Some of these enzymes have been investigated by others and us before, but sixteen target proteins represent novel sequences. Of 21 target sequences, 4 iron and 16 cobalt containing proteins were functionally expressed from BL21 (DE3) Gold. Cell free extracts were used for activity profiling and basic characterization of the NHases using the typical NHase substrate methacrylonitrile. Co-type NHases are more tolerant to high pH than Fe-type NHases. A screening for activity on three structurally diverse nitriles was carried out. Two novel Co-dependent NHases from and and a new Fe-type NHase from were very well expressed and hydrated methacrylonitrile, pyrazine-carbonitrile, and 3-amino-3-(-toluoyl)propanenitrile. The Co-dependent NHases from and , as well as two Fe-dependent NHases from , were-in addition-able to produce the amide from cinnamonitrile. Summarizing, seven so far uncharacterized NHases are described to be promising biocatalysts.
腈水合酶（NHase）催化腈转化为相应的酰胺。我们报告了各种腈水合酶的异源表达。其中一些酶以前已经被其他人以及我们研究过，但 16 个目标蛋白代表新的序列。在 21 个目标序列中，4 个含铁和 16 个含钴的蛋白从 BL21（DE3）Gold 中得到功能表达。无细胞提取物用于使用典型的 NHase 底物丙烯腈对 NHases 进行活性分析和基本特性分析。钴型 NHase 比铁型 NHase 对高 pH 值更耐受。对三种结构不同的腈进行了活性筛选。来自 和 的两种新型钴依赖型 NHase 以及来自 的新型铁依赖型 NHase 得到了很好的表达，并水合了丙烯腈、吡嗪腈和 3-氨基-3-（-甲苯酰）丙腈。来自 和 的钴依赖型 NHase 以及来自 的两种铁依赖型 NHase 除了能够从肉桂腈生成酰胺之外。总之，描述了七个迄今未表征的 NHase，它们是有前途的生物催化剂。
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