Construction of a plasmid carrying both CTP synthetase and a fused gene formed from cholinephosphate cytidylyltransferase and choline kinase genes and its application to industrial CDP-choline production: Enzymatic production of CDP-choline from orotic acid (Part II)

Article abstract of DOI:10.1271/bbb.61.960

A new method for enzymatic production of cytidine diphosphate choline (CDP-choline) from orotic acid and choline chloride was developed. To establish an industrial manufacturing process, we constructed a plasmid, pCKG55, which simultaneously expressed in Escherichia coli the three following enzymes; CTP synthetase (encoded by the pyrG gene from E. colt), cholinephosphate cytidylyltransferase (encoded by the CCT gene from Saccharomyces cerevisiae), and choline kinase (encoded by the CKI gene from S. cerevisiae). CCT and CKI genes on pCKG55 were designed to be expressed as a single CCT/CKI fused protein. This CCT/CKI fused protein retained both activities and the thermal stability of its cholinephosphate cytidylyltransferase activity was nearly the same as the native CCT enzyme. Corynebacterium ammoniagenes KY13505 and E. coli MM294/pCKG55 were cultured in 5-liter jar fermentor independently. Equal volumes of each broth were mixed in a 2-liter jar fermentor, and then the enzymatic reaction was done using 47 mM orotic acid and 60 mM choline chloride as substrates. After 23 h of the reaction at 32°C, 21.5 mM (11 g/liter) of CDP-choline was accumulated.

Full text of DOI:10.1271/bbb.61.960

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Construction of a Plasmid Carrying both CTP
Synthetase and a Fused Gene Formed from
Cholinephosphate Cytidylyltransferase and Choline
Kinase Genes and Its Application to Industrial CDP-
Choline Production: Enzymatic Production of CDP-
Choline from Orotic Acid (Part II)
Tatsuro Fujioa, Sadao Teshibab & Akihiko Maruyamab
a Planning & Development Department/Food, Liquor & Food Division, Kyowa Hakko
Kogyo Co., Ltd., 6-1 Ohtemachi, 1-chome, Chiyoda-ku, Tokyo 100, Japan
b Fine Chemicals/Biochemicals Dept., Medicals Research and Development Center,
Kyowa Hakko Kogyo Co., Ltd., 6-1 Ohtemachi, 1-chome, Chiyoda-ku, Tokyo 100, Japan
Published online: 12 Jun 2014.
To cite this article: Tatsuro Fujio, Sadao Teshiba & Akihiko Maruyama (1997) Construction of a Plasmid Carrying both
CTP Synthetase and a Fused Gene Formed from Cholinephosphate Cytidylyltransferase and Choline Kinase Genes and
Its Application to Industrial CDP-Choline Production: Enzymatic Production of CDP-Choline from Orotic Acid (Part II),
Bioscience, Biotechnology, and Biochemistry, 61:6, 960-964, DOI: 10.1271/bbb.61.960
To link to this article: http://dx.doi.org/10.1271/bbb.61.960
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