Strengthening the Ability of Pseudomonas putida to Accumulate PHA from Lignin by Metabolic Engineering

doi:10.13560/j.cnki.biotech.bull.1985.2018-0244

Abstract

Abstract:

The aim of this study is to strengthen the ability of Pseudomonas putida accumulating PHA from lignin by metabolic engineering,and to provide technical basis and research directions for improving the conversion efficiency of lignin and the production of PHA at low-cost but high-efficiency. The upstream and downstream DNA fragments of PHA helicase-encoding gene phaZ were inserted into pK18mobsacB to construct the knockout vector pK18PHAC1C2. The peroxidase-encoding gene dypB derived from R. jostii RHA1 were inserted into pVLT33 to construct the expression vector pVLTpelBDYPB. The phaZ gene was knocked out from original P. putida QSR1 with the homologous recombination,and mutant P. putida QSRZ6 was obtained. Then the peroxidase DyPB was transformed into the P. putida QSRZ6 with the heterologous recombination,P. putida QSRZ6B with DyPB expressed was constructed. The growth and PHA accumulation of mutant strains were analyzed. Cultured for 48 hours with glucose as carbon source under the N limited situation,the dry cell weight and PHA accumulation of QSRZ6 increased by 29% and 80% compared with QSR1. Cultured for 48 hours with lignin,the dry cell weight and PHA accumulation of QSRZ6 increased by 48% and 182%,as well as the PHA accumulation of P. putida QSRZ6B increased by 13% and 218% compared with the control strain P. putida QSRZ6 and P. putida QSR1,respectively,and the yield of PHA reached 140 mg/L. The ability of the strains utilizing lignin to accumulate PHA was enhanced by the phaZ-inactivation and the expression heterologous peroxidase,suggesting that metabolic engineering is an effective approach for the regulation of lignin conversion and PHA accumulation.

Key words: phaZ inactivation, lignin bioconversion, PHA accumulation, peroxidase expression, metabolic engineering

GAO Qing-long, CHEN Sheng-bao, TIAN Wen-jia, ZHANG Xue-ming, MAYu-chao. Strengthening the Ability of Pseudomonas putida to Accumulate PHA from Lignin by Metabolic Engineering[J]. Biotechnology Bulletin, 2018, 34(10): 92-99.

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