Citrulline ureidase (CTU, EC3.5.1.20) degrades citrulline into ornithine, carbon dioxide, and ammonia. Here, we present the report on expression of recombinant CTU in Escherichia coli. The soluble and active recombinant CTU was expressed in the periplasmic space with the vector pET-22b and the His-tagged CTU was purified with Ni-Affinity Chromatography. The yield of soluble recombinant protein was significantly increased when 1% sorbitol was supplemented in medium. By using phenylisothiocyanate (PITC) pre-column derivatization HPLC, the enzyme activity of recombinant CTU was determined via measuring of the substrate citrulline and the corresponding products. Our results could be useful in the study of CTU biochemical characteristics, enzymatic preparation of ornithine and development of an enzymatic detection method of citrulline.

China National Rice Research Institute Hangzhou 310006 China

College of Biological and Environmental Engineering Zhejiang University of Technology Hangzhou 310014 China

College of Chemistry and Life Sciences Zhejiang Normal University Jinhua 321004 China

Literatura

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$a Zhu, Tingheng $u College of Biological and Environmental Engineering, Zhejiang University of Technology, Hangzhou 310014, China

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$a The expression, purification and activity analysis of Francisella tularensis citrulline ureidase in Escherichia coli / $c Tingheng Zhu, Sanyu Fang, Weixia Wang, Kun Wang, Zhifeng Cui, Changchun Wang

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$a Citrulline ureidase (CTU, EC3.5.1.20) degrades citrulline into ornithine, carbon dioxide, and ammonia. Here, we present the report on expression of recombinant CTU in Escherichia coli. The soluble and active recombinant CTU was expressed in the periplasmic space with the vector pET-22b and the His-tagged CTU was purified with Ni-Affinity Chromatography. The yield of soluble recombinant protein was significantly increased when 1% sorbitol was supplemented in medium. By using phenylisothiocyanate (PITC) pre-column derivatization HPLC, the enzyme activity of recombinant CTU was determined via measuring of the substrate citrulline and the corresponding products. Our results could be useful in the study of CTU biochemical characteristics, enzymatic preparation of ornithine and development of an enzymatic detection method of citrulline.

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$a Fang, Sanyu $u College of Biological and Environmental Engineering, Zhejiang University of Technology, Hangzhou 310014, China

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$a Wang, Weixia $u China National Rice Research Institute, Hangzhou 310006, China

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$a Wang, Kun $u College of Biological and Environmental Engineering, Zhejiang University of Technology, Hangzhou 310014, China

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$a Cui, Zhifeng $u College of Biological and Environmental Engineering, Zhejiang University of Technology, Hangzhou 310014, China

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