Production of Plant Non-protein Amino Acids by Recombinant Enzymes of Sequential Biosynthetic Reactions in Bacteria
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概要
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We constructed the co-expression vector, pFK4,in which two cDNAs encoding serine acetyltransferase (SATase) and β-(pyrazol-1-yl)-L-alanine/L-cysteine synthase (β-PA/CSase) from Citrullus vulgaris (watermelon) were over-expressed under the transcriptional control of T7 promoter in Escherichia coli. Accumulation of both SATase and β-PA/CSase in soluble extracts of E. coli was confirmed by immunoblotting. The high enzymatic activities of SATase and L-cysteine synthase (CSase) were detected in cell-free extracts of E. coli carrying pFK4. The activities of the formation of β-PA and L-minosine, plant non-protein amino acids, from O-acetyl-L-serine (OAS) and the precursor heterocyclic compounds, pyrazole and 3,4-dihydroxypyridine, were also found in the extracts. β-PA was also produced in vivo from L-serine and pyrazole as precursors by E. coli cells transformed with pFK4. β-PA was accumulated mainly in the extra-cellular culture medium. The pronounced accumulation of L-cysteine and L-methionine was observed in the cells transformed with pFK4. Additionally, we also constructed vectors which carried chimeric genes encoding fusion proteins of SATase and β-PA/CSase. However, the fusion proteins tended to form insoluble inclusion bodies and thus to exhibit only weak enzymatic activities. The successful results of pFK4 shows the way to create a new sequential biosynthetic pathway of plant specific amino acids in bacterial cells by means of recombinant DNA technology.
- 公益社団法人日本薬学会の論文
- 1997-01-15
著者
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Saito K
Department Of Kampo Medicinal Science Hokkaido College Of Pharmacy
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Noji M
Laboratory Of Molecular Biology And Biotechnology Research Center Of Medicinal Resources Faculty Of
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斉藤 謙一
Department Of Physiology And Anatomy Nihon University College Of Pharmacy
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Saito K
Department Of Physiology And Anatomy Nihon University College Of Pharmacy
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SAITO Kazuki
Research Center of Medicinal Resources, Faculty of Pharmaceutical Sciences, Chiba University
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KIMURA Nobuhito
Research Center of Medicinal Resources, Faculty of Pharmaceutical Sciences, Chiba University
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IKEGAMI Fumio
Research Center of Medicinal Resources, Faculty of Pharmaceutical Sciences, Chiba University
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NOJI Masaaki
Research Center of Medicinal Resources, Faculty of Pharmaceutical Sciences, Chiba University
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Shinozaki Kazuo
Riken Kanagawa Jpn
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Ikegami Fumio
Research Center Of Medicinal Resources Faculty Of Pharmaceutical Sciences Chiba University
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Kimura Nobuhito
Department Of Molecular Biology And Biotechnology Graduate School Of Pharmaceutical Sciences Chiba U
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Kimura Nobuhito
Tokiwa Phytochemical Co.
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