Production of p‐ hydroxybenzoic acid from p‐ coumaric acid by Burkholderia glumae BGR1

ABSTRACT p ‐Coumaric acid ( p CA) is abundant in biomass with low lignin content, such as straw and stubble from rye, wheat, and barley. p CA can be isolated from biomass and used for the synthesis of aromatic hydrocarbons. Here, we report engineering of the natural pathway for conversion of p CA into p ‐hydroxybenzoic acid ( p HBA) to increase the amount of p HBA that accumulates more than 100‐fold. Burkholderia glumae strain BGR1 (BGR1) grows efficiently on p CA as a sole carbon source via a CoA‐dependent non‐β‐oxidation pathway. This pathway removes two carbons from p CA as acetyl‐CoA yielding p ‐hydroxybenzaldehyde and subsequently oxidizes it to p HBA. To increase the amount of accumulated p HBA in BGR1, we first deleted two genes encoding enzymes that degrade p HBA in the β‐ketoadipate pathway. At 10 mM of p CA, the double deletion mutant BGR1_PB4 (Δ phb3h Δ bcl ) accumulated p HBA with 95% conversion, while the control BGR1 accumulated only with 11.2% conversion. When a packed bed reactor containing immobilized BGR1_PB4 cells was operated at a dilution rate 0.2 h −1 , the productivity of p HBA was achieved at 9.27 mg/L/h for 134 h. However, in a batch reactor at 20 mM p CA, growth of BGR1_PB4 was strongly inhibited, resulting in a low conversion of 19.3%. To further increase the amount of accumulated p CA, we identified the first enzyme in the pathway, p ‐hydroxcinnmaoyl‐CoA synthetase II ( phcs II), as the rate‐limiting enzyme. Over expression of phcs II using a P alk promoter in a batch reaction at 20 mM of p CA yielded 99.0% conversion to p HBA, which is the highest concentration of p HBA ever reported using a biological process. Biotechnol. Bioeng. 2016;113: 1493–1503. © 2015 Wiley Periodicals, Inc.