Acetogenic production of 3-Hydroxybutyrate using a native 3-Hydroxybutyryl-CoA Dehydrogenase

Abstract

3-Hydroxybutyrate (3HB) is a product of interest as it is a precursor to the commercially produced bioplastic polyhydroxybutyrate. It can also serve as a platform for fine chemicals, medicines, and biofuels, making it a value-added product and feedstock. Acetogens non-photosynthetically fix CO₂ into acetyl-CoA and have been previously engineered to convert acetyl-CoA into 3HB. However, as acetogen metabolism is poorly understood, those engineering efforts have had varying levels of success. 3HB, using acetyl-CoA as a precursor, can be synthesized by a variety of different pathways. Here we systematically compare various pathways to produce 3HB in acetogens and discover a native (<i>S</i>)-3-hydroxybutyryl-CoA dehydrogenase, <i>hbd2</i>, responsible for endogenous 3HB production. In conjunction with the heterologous thiolase <i>atoB</i> and CoA transferase <i>ctfAB</i>, <i>hbd2</i> overexpression improves yields of 3HB on both sugar and syngas (CO/H₂/CO₂), outperforming the other tested pathways. These results uncovered a previously unknown 3HB production pathway, inform data from prior metabolic engineering efforts, and have implications for future physiological and biotechnological anaerobic research.