Engineering of solvent-tolerant pseudomonas putida S12 for bioproduction of phenol from glucose
article
Efficient bioconversion of glucose to phenol via the central metabolite tyrosine was achieved in the solvent-tolerant strain Pseudomonas putida S12. The tpl gene from Pantoea agglomerans, encoding tyrosine phenol lyase, was introduced into P. putida S12 to enable phenol production. Tyrosine availability was a bottleneck for efficient production. The production host was optimized by overexpressing the aroF-1 gene, which codes for the first enzyme in the tyrosine biosynthetic pathway, and by random mutagenesis procedures involving selection with the toxic antimetabolites m-fluoro-DL-phenylalanine and m-fluoro-L-tyrosine. High-throughput screening of analogue-resistant mutants obtained in this way yielded a P. putida S12 derivative capable of producing 1.5 mM phenol in a shake flask culture with a yield of 6.7% (mol/mol). In a fed-batch process, the productivity was limited by accumulation of 5 mM phenol in the medium. This toxicity was overcome by use of octanol as an extractant for phenol in a biphasic medium-octanol system. This approach resulted in accumulation of 58 mM phenol in the octanol phase, and there was a twofold increase in the overall production compared to a single-phase fed batch. Copyright © 2005, American Society for Microbiology. All Rights Reserved. Chemicals / CAS: fluorotyrosine, 139-26-4; glucose, 50-99-7, 84778-64-3; octanol, 111-87-5, 29063-28-3; phenol, 108-95-2, 3229-70-7; tyrosine, 16870-43-2, 55520-40-6, 60-18-4; DNA Primers; Glucose, 50-99-7; Phenol, 108-95-2; Tyrosine Phenol-Lyase, EC 4.1.99.2
Topics
Food technologyBioconversionEnzymesGenesGlucosePhenolsSolventsToxicityPhenol lyasePhenol productionTyrosineMicroorganisms3 fluoro dextro phenylalaninefluorotyrosineglucoseoctanolphenolphenylalanine derivativetyrosineunclassified drugbiotechnologyarticlebacterial genebacterial metabolismbioengineeringbiosynthesisbiotransformationfed batch reactorgene overexpressionhigh throughput screeningmutagenesisnonhumannucleotide sequencePantoea agglomeransPseudomonas putidatpl geneBiotransformationCloning, MolecularDNA PrimersGenetic EngineeringGlucoseKineticsMutagenesisPhenolPolymerase Chain ReactionPseudomonas putidaTyrosine Phenol-LyaseBacteria (microorganisms)Pantoea agglomeransPseudomonas putida
TNO Identifier
238851
ISSN
00992240
Source
Applied and Environmental Microbiology, 71(12), pp. 8221-8227.
Pages
8221-8227
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