TY - JOUR
T1 - Solvent resistance pumps of Pseudomonas putida S12
T2 - Applications in 1-naphthol production and biocatalyst engineering
AU - Janardhan Garikipati, S. V.B.
AU - Peeples, Tonya L.
N1 - Funding Information:
This work was done with the support of NSF grant EEC - 0310689 and the Center for Environmentally Beneficial Catalysis (CEBC) .
Publisher Copyright:
© 2015 Elsevier B.V.
PY - 2015/9/1
Y1 - 2015/9/1
N2 - The solvent resistance capacity of Pseudomonas putida S12 was applied by using the organism as a host for biocatalysis and through cloning and expressing solvent resistant pump genes into Escherichia coli. P. putida S12 expressing toluene ortho mononooxygenase (TOM-Green) was used for 1-naphthol production in a water-organic solvent biphasic system. Application of P. putida S12 improved 1-naphthol production per gram cell dry weight by approximately 42% compared to E. coli. Moreover, P. putida S12 enabled the use of a less expensive solvent, decanol, for 1-naphthol production. The solvent resistant pump (srpABC) genes of P. putida S12 were cloned into a solvent sensitive E. coli strain to transfer solvent tolerance. Recombinant strains bearing srpABC genes in either a low-copy number or a high-copy number plasmid grew in the presence of saturated concentration of toluene. Both of the recombinant strains were more tolerant to 1% v/v of toxic solvents, decanol and hexane, reaching similar cell density as the no-solvent control. Reverse-transcriptase analysis revealed that the srpABC genes were transcribed in engineered strains. The results demonstrate successful transfer of the proton-dependent solvent resistance mechanism and suggest that the engineered strain could serve as more robust biocatalysts in media with organic solvents.
AB - The solvent resistance capacity of Pseudomonas putida S12 was applied by using the organism as a host for biocatalysis and through cloning and expressing solvent resistant pump genes into Escherichia coli. P. putida S12 expressing toluene ortho mononooxygenase (TOM-Green) was used for 1-naphthol production in a water-organic solvent biphasic system. Application of P. putida S12 improved 1-naphthol production per gram cell dry weight by approximately 42% compared to E. coli. Moreover, P. putida S12 enabled the use of a less expensive solvent, decanol, for 1-naphthol production. The solvent resistant pump (srpABC) genes of P. putida S12 were cloned into a solvent sensitive E. coli strain to transfer solvent tolerance. Recombinant strains bearing srpABC genes in either a low-copy number or a high-copy number plasmid grew in the presence of saturated concentration of toluene. Both of the recombinant strains were more tolerant to 1% v/v of toxic solvents, decanol and hexane, reaching similar cell density as the no-solvent control. Reverse-transcriptase analysis revealed that the srpABC genes were transcribed in engineered strains. The results demonstrate successful transfer of the proton-dependent solvent resistance mechanism and suggest that the engineered strain could serve as more robust biocatalysts in media with organic solvents.
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U2 - 10.1016/j.jbiotec.2015.06.419
DO - 10.1016/j.jbiotec.2015.06.419
M3 - Article
C2 - 26143210
AN - SCOPUS:84937396270
SN - 0168-1656
VL - 210
SP - 91
EP - 99
JO - Journal of Biotechnology
JF - Journal of Biotechnology
ER -