High Production of 4-hydroxyvalerate by Coexpressing a 3HBDH Mutant and a NAD+-dependent Formate Dehydrogenase in Escherichia coli.

Abstract

4-Hydroxyvalerate (4HV) is an important compound that is a necessary intermediate for sustainable solvents such as gamma-valerolactone (GVL) as well as plastic additives. However, currently, the 4HV production method using the levulinic acid (LA) degradation pathway of Pseudomonas putida has a disadvantage of using a large amount of ATP. Since ATP over-consumption can affect cell growth and bio-production, it is not suitable for mass production of 4HV. Here we report alternative bio-based method of 4HV production. High production and yield of 4HV was obtained from LA, a cheap and renewable biomass source. 4HV production was performed by expressing 3-hydroxybutyrate dehydrogenase (3HBDH) that can convert LA into 4HV and formate dehydrogenase (CbFDH) that regenerates NADH in E. coli. In the first stage of cultivation, the cells were cultured at a high concentration in a pHstat-glycerol fed batch under aerobic conditions and the enzyme was expressed. In the second stage, conditions were changed to anaerobic, 4HV was produced by fed batch of LA, formate, and glycerol. As a result, about 100 g/L of 4HV was produced, which corresponds to a 95 % molar production yield. In conclusion, this conversion yield can have the potential to stimulate the development of carbon-based chemicals and promote economic access to the commercial scale of GVL.