Department of Chemistry, Hiyoshi Campus, Keio University, Kohoku-ku, Yokohama 223-8521, Japan; Laboratory of Microbial Engineering, Ōmura Satoshi Memorial Institute, Kitasato University, Sagamihara, Kanagawa 252-0373, Japan.
Laboratory of Microbial Engineering, Ōmura Satoshi Memorial Institute, Kitasato University, Sagamihara, Kanagawa 252-0373, Japan.
J Biosci Bioeng. 2020 Dec;130(6):563-570. doi: 10.1016/j.jbiosc.2020.08.006. Epub 2020 Sep 4.
To improve the production of secondary metabolites by alternation of the carbon metabolic flux, two types of deletion mutants of the central metabolic pathway, the Embden-Meyerhof (EM) or pentose phosphate (PP) pathway, in the genetically engineered Streptomyces avermitilis were constructed. Double-deletion mutants of phosphofructokinase (ΔpfkA1ΔpfkA3) in the EM pathway carrying a gene cluster for chloramphenicol biosynthesis markedly increased chloramphenicol production synthesized through the shikimate pathway. Although the ΔpfkA1ΔpfkA3 double-deletion mutant grew more slowly, its specific productivity of chloramphenicol (per dry cell weight) was 2.0-fold higher than that of the engineered S. avermitilis strain. However, the productivity of chloramphenicol was lower by the double-deletion mutant of transaldolase in the PP pathway, which supplies the precursor of the shikimate pathway. A carbon-flux analysis of the EM and PP pathways using [1-C] glucose revealed that carbon flux in the ΔpfkA1ΔpfkA3 double-deletion mutant increased through the PP pathway, which enhanced the production of chloramphenicol. These results suggest that a metabolic modification approach has the potential to increase the titers and yields of valuable secondary metabolites.
为了通过改变碳代谢通量来提高次生代谢产物的产量，构建了遗传工程化的链霉菌中两种类型的中央代谢途径（Embden-Meyerhof [EM] 或戊糖磷酸 [PP] 途径）的缺失突变体。在 EM 途径中携带氯霉素生物合成基因簇的磷酸果糖激酶（ΔpfkA1ΔpfkA3）双缺失突变体显著增加了通过莽草酸途径合成的氯霉素产量。虽然 ΔpfkA1ΔpfkA3 双缺失突变体的生长速度较慢，但氯霉素的比生产率（每干细胞重量）比工程化的链霉菌菌株高 2.0 倍。然而，PP 途径中转醛酶的双缺失突变体降低了莽草酸途径的前体供应，从而降低了氯霉素的产量。使用 [1-C] 葡萄糖对 EM 和 PP 途径的碳通量分析表明，ΔpfkA1ΔpfkA3 双缺失突变体中的碳通量通过 PP 途径增加，从而增强了氯霉素的产量。这些结果表明，代谢修饰方法有可能提高有价值的次生代谢产物的浓度和产量。
