Production of d‐p‐Hydroxyphenylglycine by N‐Carbamoyl‐d‐amino Acid Amidohydrolase‐Overproducing Escherichia coli Strains
Production of d‐p‐Hydroxyphenylglycine by N‐Carbamoyl‐d‐amino Acid Amidohydrolase‐Overproducing Escherichia coli Strains
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DOI：
10.1021/bp990060c
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发表时间：
1999
影响因子：
2.9
通讯作者：
Y. Chao;T. Juang;Jong‐Tzer Chern;Cheng-Kang Lee
中科院分区：
文献类型：
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作者：
Y. Chao;T. Juang;Jong‐Tzer Chern;Cheng-Kang Lee
从辐射农杆菌NRRL B11291中成功克隆了N-氨基甲酰-d-氨基酸酰胺水解酶（d-氨基甲酰酶）基因，并在大肠杆菌中表达。将d-氨基甲酰化酶基因亚克隆到不同类型的载体和E.大肠杆菌菌株的结果表明，在ColE 1衍生的质粒中实现了d-氨基甲酰化酶的最佳表达水平，与pSC 101衍生的质粒相比，比酶活性增加了150倍。重组质粒在大肠杆菌中的稳定性较好。coli菌株ATCC 11303，但在此处测试的JCL 1258中没有。利用重组E.大肠杆菌DH 5 α/pAH 61转化N-氨甲酰基-d-羟基苯甘氨酸为d-对羟基苯甘氨酸，转化率为100%，产率为1.9 g/（L h）。与A.辐射NRRL B11291，该生产率使d-羟基苯甘氨酸产量增加约55倍。这一结果提示了重组E. coli菌株进行转化反应。
The N‐carbamoyl‐d‐amino acid amidohydrolase (d‐carbamoylase) gene from Agrobacterium radiobacter NRRL B11291 has been successfully cloned and expressed in Escherichia coli. Subcloning of the d‐carbamoylase gene into different types of vectors and backgrounds of E. coli strains showed that the optimal expression level of d‐carbamoylase was achieved in a ColE1‐derived plasmid with a 150‐fold increase in specific enzyme activity compared to that in a pSC101‐derived plasmid. In addition, the recombinant plasmids were very stable in the E. coli strain ATCC11303 but not in JCL1258 tested here. Employing the recombinant E. coli strain DH5α/pAH61 for d‐p‐hydroxyphenylglycine production showed that the cell was capable of transforming N‐carbamoyl‐d‐hydroxylphenylglycine to d‐p‐hydroxyphenylglycine with a molar conversion yield of 100% and a production rate of 1.9 g/(L h). In comparison with A. radiobacter NRRL B11291, this productivity approximates a 55‐fold increase in d‐hydroxyphenylglycine production. This result suggests the potential application of recombinant E. coli strains for the transformation reaction.