Mechanism of adaptive laboratory evolution for improving polyunsaturated fatty acid biosynthesis in Aurantiochytrium sp. through enhancing high-glucose adaptability

Aurantiochytrium sp. (Schizochytrium sp.) is a marine fungi-like heterotrophic single-celled eukaryote utilized for the production of polyunsaturated fatty acids (PUFAs). Adaptive Laboratory Evolution (ALE) is an effective method to improve PUFAs yield. The 20th generation evolutive strain (ALE-HG20) with high PUFAs yield was screened out after high-glucose of ALE for 180 days. The yields of total lipid, eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) of ALE-HG20 strain increased by 15.51 %, 225.37 % and 52.67 % in fed-batch fermentation, respectively. Fermentation kinetics showed that the synthesis of total lipids and DHA had a tendency of uncoupling with cell growth in ALE-HG20 strain, while the synthesis of EPA had a tendency of deepening coupling with cell growth. Transcriptomic analysis revealed that the upregulation of membrane transport function genes led to rapid glucose absorption, thereby accelerating the glycolytic pathway to provide carbon skeleton for PUFAs synthesis. The metabolites of TCA pathway increased, but the amino acid anabolism associated with TCA pathway decreased, which further promoted the polysaturated fatty acid synthesis pathway. Meanwhile, the precursor metabolites of PUFAs synthesis were significantly increased. This might also be due to the protein structure evolution of GAPD and IS from glycolytic pathway and PUFAs synthesis pathway in ALE-HG20. The active pockets of GAPD and IS were increased by 10.36 % and 98.20 %, respectively, and hydrogen bonds were increased by 11.76 % and 132.76 %. This study clarified the change of regulated and evolved mechanism of PUFAs synthesis by ALE.