Citrate Synthase Overexpression of Pseudomonas putida Increases Succinate Production from Acetate in Microaerobic Cultivation.

Acetate is an end-product of anaerobic biodegradation and one of the major metabolites of microbial fermentation and lingo-cellulosic hydrolysate. Recently, acetate has been highlighted as a feedstock to produce value-added chemicals. This study examined acetate conversion to succinate by citrate synthase (gltA)-overexpressed Pseudomonas putida under microaerobic conditions. The acetate metabolism is initiated with the gltA enzyme, which converts acetyl-CoA to citrate. gltA-overexpressing P. putida (gltA-KT) showed an ∼50% improvement in succinate production compared to the wild type. Under the optimal pH of 7.5, the accumulation of succinate (4.73 ± 0.6 mM in 36 h) was ∼400% higher than that of the wild type. Overall, gltA overexpression alone resulted in 9.5% of the maximum theoretical yield in a minimal medium with acetate as the sole carbon source. This result shows that citrate synthase is important in acetate conversion to succinate by P. putida under microaerobic conditions.

Recent advances and challenges in the bioconversion of acetate to value-added chemicals.

Acetate is a major byproduct of the bioconversion of the greenhouse gas carbon dioxide, pretreatment of lignocellulose biomass, and microbial fermentation. The utilization and valorization of acetate have been emphasized in transforming waste to clean energy and value-added platform chemicals, contributing to the development of a closed carbon loop toward a low-carbon circular bio-economy. Acetate has been used to produce several platform chemicals, including succinate, 3-hydroxypropionate, and itaconic acid, highlighting the potential of acetate to synthesize many biochemicals and biofuels. On the other hand, the yields and titers have not reached the theoretical maximum. Recently, recombinant strain development and pathway regulation have been suggested to overcome this limitation. This review provides insights into the important constraints limiting the yields and titers of the biochemical and metabolic pathways of bacteria capable of metabolizing acetate for acetate bioconversion. The current developments in recombinant strain engineering are also discussed.

Colorimetric isolation of a novel electrochemically active Pseudomonas strain using tungsten nanorods for bioelectrochemical applications.

Pseudomonas strains are a promising host cell in metabolic engineering for bioconversion, environmental remediation, and most recently for bioelectrochemical applications. This study isolated an electrochemically active Pseudomonas sp. from an anaerobic sludge using a colorimetric and electrochromic WO3 nanorod (WO3-NR) probe. A strategy was developed to determine the presence of electroactive species from enriched cultures. A mixed consortium was enriched using Pseudomonas isolation media containing betaine and triclosan as the carbon source and antibacterial reagent, respectively. A single blue colony was isolated using WO3-NR sandwiched agar plates. The isolate was sequenced by 16 s rRNA and designated Pseudomonas aeruginosa PBH03, producing phenazines and pyocyanin aerobically. The isolate exhibited clear electrochemical characteristics from cyclic voltammetry and linear sweep voltammetry and produced a current density of 9.01 µA cm-2 in a microbial fuel cell.

Enabling anoxic acetate assimilation by electrode-driven respiration in the obligate aerobe, Pseudomonas putida.

This study examined the obligate aerobe, Pseudomonas putida, using acetate as the sole carbon and energy source, and respiration via an anode as the terminal electron acceptor under anoxic conditions. P. putida showed significantly different acetate assimilation in a closed-circuit microbial fuel cell (CC-MFC) compared to an open circuit MFC (OC-MFC). More than 72% (2.6 mmol) of acetate was consumed during 84 hrs in the CC-MFC in contrast to the no acetate consumption observed in the OC-MFC. The CC-MFC produced 150 µA (87 C) from acetate metabolization. Electrode-based respiration reduced the NADH/NAD+ ratio anaerobically, which is similar to the aerobic condition. The CC-MFC showed significantly higher acetyl-CoA synthetase activity than the OC-MFC (0.028 vs. 0.001 µmol/min/mg), which was comparable to the aerobic condition (circa 60%). Overall, electrode-based respiration enables P. putida to metabolize acetate under anoxic conditions and provide a platform to regulate the bacterial redox balance without oxygen.