Efficient Production of 2,4-Dihydroxybutyrate from l-Homoserine by the Designed Cofactor Self-Sufficient Route

2,4-Dihydroxybutyric acid (DHB) is an α-hydroxy acid that has the potential as a versatile precursor. As a starting material for the chemical synthesis of the l-methionine analogue, 2-hydroxy-4-(methylthio)butyrate, which targets a valuable market in animal nutrition, has made DHB gain considerable attention. In this study, we designed a cofactor self-sufficient route for DHB production from l-homoserine by introducing glutamate dehydrogenase to synergistically regenerate cosubstrate and maintain redox balance, which significantly boosted DHB production from l-homoserine. The efficiency was further improved by screening for new dehydrogenases with high 2-keto-4-hydroxybutyrate reductase activity and optimizing enzyme expression. The engineered strain could produce 78.8 mM DHB from 83.5 mM l-homoserine with a molar yield of 94.4%. In addition, an equivalent yield with a high titer (148.2 mM) was achieved in the sequential fermentation–biotransformation process to produce DHB directly from the fermentation broth containing l-homoserine. Thus, this novel design of constructing a cofactor self-circulation whole-cell biocatalytic system provided an efficient alternative for synthesizing DHB from renewable feedstocks.