Abstract
The bacteria that grow on methane aerobically (methanotrophs) support populations of non-methanotrophs in the natural environment by excreting methane-derived carbon. One group of excreted compounds are short-chain organic acids, generated in highest abundance when cultures are grown under O2-starvation. We examined this O2-starvation condition in the methanotroph Methylomicrobium buryatense 5GB1. The M. buryatense 5GB1 genome contains homologs for all enzymes necessary for a fermentative metabolism, and we hypothesize that a metabolic switch to fermentation can be induced by low-O2 conditions. Under prolonged O2-starvation in a closed vial, this methanotroph increases the amount of acetate excreted about 10-fold, but the formate, lactate, and succinate excreted do not respond to this culture condition. In bioreactor cultures, the amount of each excreted product is similar across a range of growth rates and limiting substrates, including O2-limitation. A set of mutants were generated in genes predicted to be involved in generating or regulating excretion of these compounds and tested for growth defects, and changes in excretion products. The phenotypes and associated metabolic flux modeling suggested that in M. buryatense 5GB1, formate and acetate are excreted in response to redox imbalance. Our results indicate that even under O2-starvation conditions, M. buryatense 5GB1 maintains a metabolic state representing a combination of fermentation and respiration metabolism.
Highlights
Methanotrophs are a group of bacteria able to grow on methane as sole carbon and energy source (Whittenbury, Phillips & Wilkinson, 1970)
It has been shown that a gamma-proteobacterial methanotroph, Methylomicrobium alcaliphilum 20Z, contains a highly efficient version of the ribulose monophosphate cycle for formaldehyde assimilation, that could theoretically allow for a fermentation type of metabolism, with O2 used for activating the methane molecule, but not as a terminal electron acceptor (Kalyuzhnaya et al, 2013)
Strains tested for these excretion products so far are both haloalkaliphiles (Kaluzhnaya et al, 2001), we assessed two different gamma-proteobacterial methanotrophs isolated from a freshwater lake, Methylobacter tundripaludum 31/32 and Methylomonas strain LW13 (Kalyuzhnaya et al, 2015), and showed that these strains generate formate, acetate, and succinate under these growth conditions, but lactate and hydroxybutyrate were not detected (Fig. 2)
Summary
Methanotrophs are a group of bacteria able to grow on methane as sole carbon and energy source (Whittenbury, Phillips & Wilkinson, 1970) They play an important role in natural habitats, retaining methane carbon in ecosystems and mitigating emissions of this potent greenhouse gas (Knief, 2015; Chistoserdova, 2015). Evidence was provided that genes encoding enzymes of such a pathway are widespread in gamma-proteobacterial methanotrophs, and are transcriptionally up-regulated when cells are cultured under O2-starvation conditions, concomitant with excretion of putative fermentation end products (Kalyuzhnaya et al, 2013) These results are intriguing, both for the potential of methanotrophs to cross-feed non-methanotrophs with methane-derived carbon in natural communities (Radajewski, McDonald & Murrell, 2003; Oshkin et al, 2015), and for the potential to manipulate methanotrophic metabolism to generate excreted products (Kalyuzhnaya, Puri & Lidstrom, 2015)
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