LysR-type transcriptional regulator TtdR regulates both ethylene glycol and polyhydroxyalkanoate (PHA) metabolism in Pseudomonas umsongensis GO16

Abstract In Pseudomonas umsongensis GO16, a LysR-type transcriptional regulator (LTTRs) ttdR is located 5′ of the gene for glyoxylate carboligase ( gcl ), involved in ethylene glycol (EG) metabolism. GO16 wildtype (WT) can utilize EG as a sole carbon and energy source but the ttdR knockout strain (Δ ttdR ) cannot grow on EG, demonstrating its role as an activator in EG metabolism. When Δ ttdR was grown with terephthalic acid (TA) or glucose, the growth was comparable to the WT. GO16 originally produces both C4 short-chain-length (scl) PHA and C6-C12 medium-chain-length (mcl) PHA from various carbon sources. Interestingly, Δ ttdR produced scl-PHA monomer 5.4-fold and 1.4-fold increased than the WT when TA and glucose were used respectively. Furthermore, Δ ttdR exhibited a very long lag phase when grown with fatty acids, namely butyrate or octanoate. This indicates a more complex role of TtdR than simply activating EG metabolism in GO16. The analysis of the GO16 Δ ttdR proteome revealed that EG catabolic genes were expressed, but their abundance was 2.8- to 10.5-fold lower (log 2 ) compared to the WT. In addition, higher abundance of enzymes that could be contributing to higher scl-PHA content was also observed. GO16 Δ ttdR was further subjected to adaptive laboratory evolution (ALE) with butyrate and octanoate. The evolved strains regained the ability to grow with these fatty acids as the WT, but the mutations accumulated did not confer growth with EG or acetate, suggesting that a mechanism different to glyoxylate shunt has evolved to allow growth with fatty acids.