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Increasing pinosylvin production in Escherichia coli by reducing the expression level of the gene fabI-encoded enoyl-acyl carrier protein reductase
Salas-Navarrete, Caheri; Hernández-Chávez, Georgina; Flores, Noemí; Martínez, Luz María; Martinez, Alfredo; Bolívar, Francisco; Barona-Gomez, Francisco; Gosset, Guillermo.
  • Salas-Navarrete, Caheri; Universidad Nacional Autónoma de México. Departamento de Ingeniería Celular y Biocatálisis. Instituto de Biotecnología. Cuernavaca. MX
  • Hernández-Chávez, Georgina; Universidad Nacional Autónoma de México. Departamento de Ingeniería Celular y Biocatálisis. Instituto de Biotecnología. Cuernavaca. MX
  • Flores, Noemí; Universidad Nacional Autónoma de México. Departamento de Ingeniería Celular y Biocatálisis. Instituto de Biotecnología. Cuernavaca. MX
  • Martínez, Luz María; Universidad Nacional Autónoma de México. Departamento de Ingeniería Celular y Biocatálisis. Instituto de Biotecnología. Cuernavaca. MX
  • Martinez, Alfredo; Universidad Nacional Autónoma de México. Departamento de Ingeniería Celular y Biocatálisis. Instituto de Biotecnología. Cuernavaca. MX
  • Bolívar, Francisco; Universidad Nacional Autónoma de México. Departamento de Ingeniería Celular y Biocatálisis. Instituto de Biotecnología. Cuernavaca. MX
  • Barona-Gomez, Francisco; Laboratorio Nacional de Genómica para la Biodiversidad, Cinvestav ­ IPN. Irapuato. MX
  • Gosset, Guillermo; Universidad Nacional Autónoma de México. Departamento de Ingeniería Celular y Biocatálisis. Instituto de Biotecnología. Cuernavaca. MX
Electron. j. biotechnol ; 33: 11-16, May. 2018. ilus, tab, graf
Article in English | LILACS | ID: biblio-1022822
ABSTRACT

Background:

The plant secondary metabolite pinosylvin is a polyphenol from the stilbene family, which have positive effects on human health. Biotechnological production is an attractive alternative for obtaining this stilbene. In Escherichia coli, malonyl-CoA is the precursor for both stilbene and fatty acid syntheses. In this study, with the aim of increasing pinosylvin production, we evaluated a novel approach that is based on reducing the expression of the gene fabI, which encodes the enzyme enoyl-acyl carrier protein reductase that is involved in fatty acid synthesis.

Results:

A recombineering method was employed to eliminate the chromosomal -35 promoter sequence and the upstream region of the gene fabI in E. coli strain W3110. Analysis, employing RT-qPCR, showed that such modification caused a 60% reduction in the fabI transcript level in the mutant strain W3110Δ-35fabICm compared to the wild type W3110. Synthetic genes encoding a mutant version of 4-coumaroyl-CoA ligase from Streptomyces coelicolor A3 with improved catalytic activity employing cinnamic acid as substrate and a stilbene synthase from Vitis vinifera were cloned to generate the plasmid pTrc-Sc4CL(M)-VvSTS. The production performance of strains W3110Δ-35fabICm/pTrc-Sc4CL(M)-VvSTS and W3110/pTrc-Sc4CL(M)- VvSTS was determined in shake flask cultures with Luria-Bertani medium supplemented with 10 g/L glycerol and 3 mM cinnamic acid. Under these conditions, the strain W3110Δ-35fabICm/pTrc-Sc4CL(M)-VvSTS produced 52.67 mg/L pinosylvin, a level 1.5-fold higher than that observed with W3110/pTrc-Sc4CL(M)-VvSTS.

Conclusion:

A reduction in the transcript level of fabI caused by the elimination of the -35 and upstream promoter sequences is a successful strategy to improve pinosylvin production in E. coli.
Subject(s)


Full text: Available Index: LILACS (Americas) Main subject: Stilbenes / Escherichia coli / Enoyl-(Acyl-Carrier-Protein) Reductase (NADH) Language: English Journal: Electron. j. biotechnol Journal subject: Biotechnology Year: 2018 Type: Article Affiliation country: Mexico Institution/Affiliation country: Laboratorio Nacional de Genómica para la Biodiversidad, Cinvestav ­ IPN/MX / Universidad Nacional Autónoma de México/MX

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Full text: Available Index: LILACS (Americas) Main subject: Stilbenes / Escherichia coli / Enoyl-(Acyl-Carrier-Protein) Reductase (NADH) Language: English Journal: Electron. j. biotechnol Journal subject: Biotechnology Year: 2018 Type: Article Affiliation country: Mexico Institution/Affiliation country: Laboratorio Nacional de Genómica para la Biodiversidad, Cinvestav ­ IPN/MX / Universidad Nacional Autónoma de México/MX