[Cloning and expression of variants of the glutaryl-7-aminocephalosporic acid acylase of the bacterium Brevundimonas diminuta in Escherichia coli cells]статья
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Аннотация:The gene coding for glutaryl-7-aminocephalosporic acid acylase (Gl7ACA acylase) of the bacterium Brevundimonas diminuta (BrdGl7ACA), a commercial enzyme widely used in modem biocatalytic technologies for manufacture of b-lactam antibiotics, was cloned. Efficient expression systems for producing a "native" recombinant BrdGl7ACA and its analogs modified by attaching affinity groups–the chitin-binding domain of chitinases A1 and hexahistidine sequence–were designed. It was demonstrated that both the recombinant hybrid proteins and the native Gl7ACA acylase produced in E. coli cells underwent a correct autoproteolytic processing with generation of functionally active enzymes and could be isolated with a high yield using one-step affinity chromatography.