Metabolic engineering of genome-streamlined strain Pseudomonas putida KTU-U27 for medium-chain-length polyhydroxyalkanoate production from xylose and cellobiose

Honglu Liu; Yaping Chen; Siqi Wang; Yujie Liu; Wanwan Zhao; Kaiyue Huo; Hongfu Guo; Weini Xiong; Shufang Wang; Chao Yang; Ruihua Liu

doi:10.1016/j.ijbiomac.2023.126732

Metabolic engineering of genome-streamlined strain Pseudomonas putida KTU-U27 for medium-chain-length polyhydroxyalkanoate production from xylose and cellobiose

Int J Biol Macromol. 2023 Dec 31;253(Pt 2):126732. doi: 10.1016/j.ijbiomac.2023.126732. Epub 2023 Sep 5.

Authors

Honglu Liu¹, Yaping Chen¹, Siqi Wang¹, Yujie Liu¹, Wanwan Zhao¹, Kaiyue Huo¹, Hongfu Guo¹, Weini Xiong¹, Shufang Wang², Chao Yang³, Ruihua Liu⁴

Affiliations

¹ Key Laboratory of Molecular Microbiology and Technology for Ministry of Education, College of Life Sciences, Nankai University, Tianjin 300071, China.
² Key Laboratory of Bioactive Materials for Ministry of Education, College of Life Sciences, Nankai University, Tianjin 300071, China. Electronic address: wangshufang@nankai.edu.cn.
³ Key Laboratory of Molecular Microbiology and Technology for Ministry of Education, College of Life Sciences, Nankai University, Tianjin 300071, China. Electronic address: yangc20119@nankai.edu.cn.
⁴ Tianjin Key Laboratory of Protein Science, College of Life Sciences, Nankai University, Tianjin 300071, China. Electronic address: yangyangliu@nankai.edu.cn.

PMID: 37678685
DOI: 10.1016/j.ijbiomac.2023.126732

Abstract

Bio-based plastics polyhydroxyalkanoates (PHAs) are considered as a good substitutive to traditional fossil-based plastics because PHAs outcompete chemical plastics in several important properties, such as biodegradability, biocompatibility, and renewability. However, the industrial production of PHA (especially medium-chain-length PHA, mcl-PHA) is greatly restricted by the cost of carbon sources. Currently, xylose and cellobiose derived from lignocellulose are potential substrates for mcl-PHA production. In this study, Pseudomonas putida KTU-U27, a genome-streamlined strain derived from a mcl-PHA producer P. putida KT2440, was used as the optimal chassis for the construction of microbial cell factories with the capacity to efficiently produce mcl-PHA from xylose and cellobiose by introducing the xylose and cellobiose metabolism modules and enhancing the transport of xylose and cellobiose. The lag phases of the xylose- and cellobiose-grown engineered strains were almost completely eliminated and the xylose- and cellobiose-utilizing performance was greatly improved via adaptive laboratory evolution. In shake-flask fermentation, the engineered strain 27A-P13-xylABE-P_tac-tt and 27A-P13-bglC-P13-gts had a mcl-PHA content of 41.67 wt% and 45.18 wt%, respectively, and were able to efficiently utilize xylose or cellobiose as the sole carbon source for cell growth. Herein, microbial production of mcl-PHA using xylose as the sole carbon source has been demonstrated for the first time. Meanwhile, the highest yield of mcl-PHA produced from cellobiose has been obtained in this study. Interestingly, the engineered strains derived from genome-reduced P. putida strains showed higher xylose- and cellobiose-utilizing performance and higher PHA yield than those derived from P. putida KT2440. This study highlights enormous potential of the engineered strains as promising platforms for low-cost production of mcl-PHA from xylose- and cellobiose-rich substrates.

Keywords: Cellobiose; Genome-streamlined strain; Minimal genome cell factories; Polyhydroxyalkanoate; Pseudomonas putida KT2440; Xylose.

MeSH terms

Carbon / metabolism
Cellobiose / metabolism
Metabolic Engineering
Polyhydroxyalkanoates*
Pseudomonas putida* / genetics
Pseudomonas putida* / metabolism
Xylose / metabolism

Substances

Xylose
Cellobiose
Polyhydroxyalkanoates
Carbon