Production of nonproteinaceous amino acids using recombinant Escherichia coli cells expressing cysteine synthase and related enzymes with or without the secretion of O-acetyl-L-serine

Chunhui Zhao, Katsuhiro Ohno, Kohji Sogoh, Koreyoshi Imamura, Takaharu Sakiyama, Kazuhiro Nakanishi

Research output: Contribution to journalArticle

8 Citations (Scopus)

Abstract

β-(Pyrazol-1-yl)-L-alanine (β-PA), a model nonproteinaceous amino acid, was specifically synthesized by two methods using recombinant Escherichia coli cells that express cysteine syathase, comprising serine acetyltransferase (SAT) and O-acetylserine sulfhydrylase-A (OASS-A) and related enzymes firooi E. coli. In the first method (method A), recombinant cells that express wildtype SAT, OASS-A, acetate kinase (AK), and phosphotransacetylase (PTA) showed the highest β-PA production. β-PA was produced at 140 mM from 200 mM L-serine and 200 mM pyrazole under optimum conditions. Using the cells expressing SATΔC20 (truncated SAT), OASS-A, AK, and PTA, β-PA was produced at a level of only 80 mM, whereas O-acetyl-serine (OAS) was found to be secreted into the broth. Under optimum conditions, OAS accumulated at levels of around 105 mM from 300 mM L-serine. Thus, in the second method (method B), the secreted OAS was used as the substrate for the syntheses of β-PA and β-(triazol-1-yl)-L-alanine (β-TA). The OAS that accumulated in the broth was efficiently converted to β-PA and β-TA at levels of around 90 mM from 105 mM OAS using free OASS-A. In both methods A and B, the addition of glucose was essential for the efficient production of β-PA and OAS, respectively.

Original languageEnglish
Pages (from-to)322-328
Number of pages7
JournalJournal of Bioscience and Bioengineering
Volume97
Issue number5
Publication statusPublished - 2004

Fingerprint

Cysteine Synthase
Serine
Escherichia coli
Amino acids
Enzymes
Cells
Serine O-Acetyltransferase
Amino Acids
Glucose
Phosphate Acetyltransferase
Acetate Kinase
Substrates
Alanine
Cysteine

Keywords

  • β-(pyrazol-1-yl)- L-alanine
  • Cysteine synthase
  • Escherichia coli
  • Nonproteinaceous amino acid
  • O-acetyl-L-serine

ASJC Scopus subject areas

  • Biotechnology
  • Bioengineering

Cite this

Production of nonproteinaceous amino acids using recombinant Escherichia coli cells expressing cysteine synthase and related enzymes with or without the secretion of O-acetyl-L-serine. / Zhao, Chunhui; Ohno, Katsuhiro; Sogoh, Kohji; Imamura, Koreyoshi; Sakiyama, Takaharu; Nakanishi, Kazuhiro.

In: Journal of Bioscience and Bioengineering, Vol. 97, No. 5, 2004, p. 322-328.

Research output: Contribution to journalArticle

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abstract = "β-(Pyrazol-1-yl)-L-alanine (β-PA), a model nonproteinaceous amino acid, was specifically synthesized by two methods using recombinant Escherichia coli cells that express cysteine syathase, comprising serine acetyltransferase (SAT) and O-acetylserine sulfhydrylase-A (OASS-A) and related enzymes firooi E. coli. In the first method (method A), recombinant cells that express wildtype SAT, OASS-A, acetate kinase (AK), and phosphotransacetylase (PTA) showed the highest β-PA production. β-PA was produced at 140 mM from 200 mM L-serine and 200 mM pyrazole under optimum conditions. Using the cells expressing SATΔC20 (truncated SAT), OASS-A, AK, and PTA, β-PA was produced at a level of only 80 mM, whereas O-acetyl-serine (OAS) was found to be secreted into the broth. Under optimum conditions, OAS accumulated at levels of around 105 mM from 300 mM L-serine. Thus, in the second method (method B), the secreted OAS was used as the substrate for the syntheses of β-PA and β-(triazol-1-yl)-L-alanine (β-TA). The OAS that accumulated in the broth was efficiently converted to β-PA and β-TA at levels of around 90 mM from 105 mM OAS using free OASS-A. In both methods A and B, the addition of glucose was essential for the efficient production of β-PA and OAS, respectively.",
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AU - Zhao, Chunhui

AU - Ohno, Katsuhiro

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AU - Imamura, Koreyoshi

AU - Sakiyama, Takaharu

AU - Nakanishi, Kazuhiro

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N2 - β-(Pyrazol-1-yl)-L-alanine (β-PA), a model nonproteinaceous amino acid, was specifically synthesized by two methods using recombinant Escherichia coli cells that express cysteine syathase, comprising serine acetyltransferase (SAT) and O-acetylserine sulfhydrylase-A (OASS-A) and related enzymes firooi E. coli. In the first method (method A), recombinant cells that express wildtype SAT, OASS-A, acetate kinase (AK), and phosphotransacetylase (PTA) showed the highest β-PA production. β-PA was produced at 140 mM from 200 mM L-serine and 200 mM pyrazole under optimum conditions. Using the cells expressing SATΔC20 (truncated SAT), OASS-A, AK, and PTA, β-PA was produced at a level of only 80 mM, whereas O-acetyl-serine (OAS) was found to be secreted into the broth. Under optimum conditions, OAS accumulated at levels of around 105 mM from 300 mM L-serine. Thus, in the second method (method B), the secreted OAS was used as the substrate for the syntheses of β-PA and β-(triazol-1-yl)-L-alanine (β-TA). The OAS that accumulated in the broth was efficiently converted to β-PA and β-TA at levels of around 90 mM from 105 mM OAS using free OASS-A. In both methods A and B, the addition of glucose was essential for the efficient production of β-PA and OAS, respectively.

AB - β-(Pyrazol-1-yl)-L-alanine (β-PA), a model nonproteinaceous amino acid, was specifically synthesized by two methods using recombinant Escherichia coli cells that express cysteine syathase, comprising serine acetyltransferase (SAT) and O-acetylserine sulfhydrylase-A (OASS-A) and related enzymes firooi E. coli. In the first method (method A), recombinant cells that express wildtype SAT, OASS-A, acetate kinase (AK), and phosphotransacetylase (PTA) showed the highest β-PA production. β-PA was produced at 140 mM from 200 mM L-serine and 200 mM pyrazole under optimum conditions. Using the cells expressing SATΔC20 (truncated SAT), OASS-A, AK, and PTA, β-PA was produced at a level of only 80 mM, whereas O-acetyl-serine (OAS) was found to be secreted into the broth. Under optimum conditions, OAS accumulated at levels of around 105 mM from 300 mM L-serine. Thus, in the second method (method B), the secreted OAS was used as the substrate for the syntheses of β-PA and β-(triazol-1-yl)-L-alanine (β-TA). The OAS that accumulated in the broth was efficiently converted to β-PA and β-TA at levels of around 90 mM from 105 mM OAS using free OASS-A. In both methods A and B, the addition of glucose was essential for the efficient production of β-PA and OAS, respectively.

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