Electron availability in CO2 , CO and H2 mixtures constrains flux distribution, energy management and product formation in Clostridium ljungdahlii

Abstract

Acetogens such as Clostridium ljungdahlii can play a crucial role reducing the human CO₂ footprint by converting industrial emissions containing CO₂ , CO and H₂ into valuable products such as organic acids or alcohols. The quantitative understanding of cellular metabolism is a prerequisite to exploit the bacterial endowments and to fine-tune the cells by applying metabolic engineering tools. Studying the three gas mixtures CO₂ + H₂ , CO and CO + CO₂ + H₂ (syngas) by continuously gassed batch cultivation experiments and applying flux balance analysis, we identified CO as the preferred carbon and electron source for growth and producing alcohols. However, the total yield of moles of carbon (mol-C) per electrons consumed was almost identical in all setups which underlines electron availability as the main factor influencing product formation. The Wood-Ljungdahl pathway (WLP) showed high flexibility by serving as the key NAD⁺ provider for CO₂ + H_2, whereas this function was strongly compensated by the transhydrogenase-like Nfn complex when CO was metabolized. Availability of reduced ferredoxin (Fd_red ) can be considered as a key determinant of metabolic control. Oxidation of CO via carbon monoxide dehydrogenase (CODH) is the main route of Fd_red formation when CO is used as substrate, whereas Fd_red is mainly regenerated via the methyl branch of WLP and the Nfn complex utilizing CO₂ + H₂ . Consequently, doubled growth rates, highest ATP formation rates and highest amounts of reduced products (ethanol, 2,3-butanediol) were observed when CO was the sole carbon and electron source.