| PHAs is a kind of biocompatible and biodegradable biopolyester accumulated by microorganisms in the form of small particles.It is an ideal biological material to replace traditional plastics and alleviate the environmental problems caused by the accumulation of non-degradable traditional plastic.P(3HB-co-LA)belongs to PHAs,compared to P(3HB)and PLA,it has better toughness and transparency.In this study,β-ketothiolase(phaA),acetoacetyl-CoA reductase(phaB)and polyhydroxyalkanoate synthase(phaCm)required for PHAs synthesis and mutanted propionate coenzyme A transferase(pctth)were introduced into E.coli MG1655 to synthesize P(3HB-co-LA).In order to increase the lactate fraction in P(3HB-co-LA),the gene D-lactate dehydrogenase(dld)was knocked out.In the optimized xylose fermentation,the lactate fraction in P(3HB-co-LA)synthesized by the dld deficient strain WXJ01-03 has increased from 23.8 mol%to 37.2 mol%.The endogenous thioesterase of E.coli can degrade the acyl-CoA,and the thioesterase ydiI and yciA with degrading effect on LA-CoA were screened in this study.Two thioesterases were knocked out based on strain WXJ01-03 to obtain the recombinant strainWXJ03-03.Finally,the lactate fraction in P(3HB-co-LA)synthesized by WXJ03-03 reached 46.1 mol%in xylose fermentation.This paper also tries using glucose instead of xylose to produce P(3HB-co-LA),results suggest that when glucose as carbon source,the strains grow faster during the early stage of fermentation and the overall carbon source consumption is less than xylose.However the lack of thioesterase in E.coli can not increase lactate fraction in co-polymer.Transcriptional data showed that the key gene(pctth)was poorly transcribed when take glucose as carbon source,thus lack of thioesterase could hardly improve the lactate fraction of P(3HB-co-LA).In order to comprehensively combine the advantages of glucose and xylose fermentation,mixed sugar(a mixture of glucose and xylose)was used in this study to produce P(3HB-co-LA),it also provide a theoretical basis for the biosynthesis of bio-based polymers by using cheap and easily available lignin hydrolysate as carbon source in the future. |
