Ferredoxin-dependent metabolic engineering of electron transfer circuits has been developed to enhance redox efficiency in the field of synthetic biology, e.g., for hydrogen production and for reduction of flavoproteins or NAD(P)+. Here, we present the bioconversion of carbon monoxide (CO) gas to formate via a synthetic CO:formate oxidoreductase (CFOR), designed as an enzyme complex for direct electron transfer between non-interacting CO dehydrogenase and formate dehydrogenase using an electron-transferring Fe-S fusion protein. The CFOR-introduced Thermococcus onnurineus mutant strains showed CO-dependent formate production in vivo and in vitro. The maximum formate production rate from purified CFOR complex and specific formate productivity from the bioreactor were 2.2 ± 0.2 μmol/mg/min and 73.1 ± 29.0 mmol/g-cells/h, respectively. The CO-dependent CO2 reduction/formate production activity of synthetic CFOR was confirmed, indicating that direct electron transfer between two unrelated dehydrogenases was feasible via mediation of the FeS-FeS fusion protein.

The genome of the hyperthermophilic archaeon Thermococcus onnurineus NA1 contains three copies of the formate dehydrogenase (FDH) gene, fdh1, fdh2, and fdh3. Previously, we reported that fdh2, clustered with genes encoding the multimeric membrane-bound hydrogenase and cation/proton antiporter, was essential for formate-dependent growth with H2 production. However, the functionality of the other two FDH-coding genes has not yet been elucidated. Herein, we purified and characterized cytoplasmic Fdh3 to understand its functionality. The purified Fdh3 was identified to be composed of a tungsten-containing catalytic subunit (Fdh3A), an NAD(P)-binding protein (Fdh3B), and two Fe-S proteins (Fdh3G1 and Fdh3G2). Fdh3 oxidized formate with specific activities of 241.7 U/mg and 77.4 U/mg using methyl viologen and NADP+ as electron acceptors, respectively. While most FDHs exhibited NAD+-dependent formate oxidation activity, the Fdh3 of T. onnurineus NA1 showed a strong preference for NADP+ over NAD+ as a cofactor. The catalytic efficiency (kcat/Km) of Fdh3 for NADP+ was measured to be 5,281 mM−1 s−1, which is the highest among NADP-dependent FDHs known to date. Structural modeling suggested that Arg204 and Arg205 of Fdh3B may contribute to the stabilization of the 2′-phosphate of NADP(H). Fdh3 could also use ferredoxin as an electron acceptor to oxidize formate with a specific activity of 0.83 U/mg. Furthermore, Fdh3 showed CO2 reduction activity using reduced ferredoxin or NADPH as an electron donor with a specific activity of 0.73 U/mg and 1.0 U/mg, respectively. These results suggest a functional role of Fdh3 in disposing of reducing equivalents by mediating electron transfer between formate and NAD(P)H or ferredoxin.

Background: This study aimed to identify the incidence rate of post-coronavirus disease-2019 (COVID-19) conditions in the Republic of Korea (ROK) Army and to investigate the trend of the incidence rate according to changes in dominant variants.Methods: We used the results of a 19-item, self-completed survey of those who had recovered from COVID-19 in the ROK Army between March 24, 2020, and April 30, 2022. We used both descriptive and multiple logistic regression analyses to identify factors associated with the incidence rate of post-COVID-19 conditions.Results: Among the total of 48,623 COVID-19 episodes in the ROK Army, the overall incidence rate of post-COVID-19 conditions was 32.9%. Based on the survey, the incidence of cough was the highest at 15.4%, followed by fatigue (15.1%) and sputum (13.8%). The delta variant had the highest incidence rate of post-COVID-19 conditions at 50.7%, whereas the omicron variant had the lowest at 19.7%. Concerning the type of post-COVID-19 condition, the neuropsychiatric symptoms had the highest incidence at 27.4% when the delta variant was dominant, and the respiratory symptoms were highest at 37.3% when the omicron variant was dominant. In the case of smell and taste symptoms, the incidence rate was high at 21.1% only when the delta variant was predominant.Conclusion: The overall incidence rate of post-COVID-19 conditions in the ROK Army was 32.9%. When the delta variant was dominant, the overall incidence as well as the proportion of neuropsychiatric symptoms were high. However, as the omicron variant became dominant, the overall incidence decreased, but the proportion of respiratory symptoms increased.