期刊
METABOLIC ENGINEERING
卷 66, 期 -, 页码 229-238出版社
ACADEMIC PRESS INC ELSEVIER SCIENCE
DOI: 10.1016/j.ymben.2021.04.015
关键词
Para-coumarate; Bioreactor; RB-TnSeq; Pseudomonas putida; Metabolic engineering
资金
- US Department of Energy, Office of Science [DE-AC02-05CH11231]
- US Department of Energy, BioEnergy Technology Office
Pseudomonas putida KT2440, manipulated through disrupting a majority of non-essential genes, showed improved fitness in stirred-tank bioreactor, resulting in enhanced production of indigoidine from para-coumarate, indicating its potential as a promising biomanufacturing host for the conversion of renewable carbon streams.
Pseudomonas putida KT2440 is an emerging biomanufacturing host amenable for use with renewable carbon streams including aromatics such as para-coumarate. We used a pooled transposon library disrupting nearly all (4,778) non-essential genes to characterize this microbe under common stirred-tank bioreactor parameters with quantitative fitness assays. Assessing differential fitness values by monitoring changes in mutant strain abundance identified 33 gene mutants with improved fitness across multiple stirred-tank bioreactor formats. Twentyone deletion strains from this subset were reconstructed, including GacA, a regulator, TtgB, an ABC transporter, and PP_0063, a lipid A acyltransferase. Thirteen deletion strains with roles in varying cellular functions were evaluated for conversion of para-coumarate, to a heterologous bioproduct, indigoidine. Several mutants, such as the Delta gacA strain improved fitness in a bioreactor by 35 fold and showed an 8-fold improvement in indigoidine production (4.5 g/L, 0.29 g/g, 23% of maximum theoretical yield) from para-coumarate as the carbon source.
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