Methylomonas methanica and Methylosinus sp. co-cultivated with methylotrophs under saline conditions for microbial protein production from biogas as alternatives to natural-gas- based production using Methylococcus capsulatus

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Abstract
Introduction: Microbial protein (MP) is a promising alternative protein source that can be produced using methane as a substrate derived from undervalued resources. Methylococcus capsulatus is one of the most prominent microbial species employed for industrial production, while other potentially suitable methanotrophs remain underexplored. Methods: Here, we selected the alternative methanotrophs Methylosinus sp. and Methylomonas methanica with high growth rates and potential for halotolerance and compared them to M. capsulatus . Results and Discussion: Methylosinus sp. and Methylomonas methanica achieved up to 1.87-fold higher maximum specific growth rates and 2.52-fold higher biomass concentrations, compared to M. capsulatus . When the pH was allowed to acidify below optimum for growth, the protein content of M. methanica (45% ± 5% CDW) was 1.55 times compared to pH-controlled conditions. Cultivation at an increased salt content (20 g NaCl L −1 ) boosted the protein content of M. methanica to 81% ± 7% CDW, showing its applicability for MP production using saline water. Co-culturing M. methanica with the methylotrophic yeast Komagataella phaffii led to a 1.27-fold higher essential amino acid content, which indicates an improvement in protein quality, while the use of synthetic biogas increased the protein yields up to 1.5-fold, compared to purified methane. In summary, M. methanica co-cultured with selected methylotrophs has the potential to enable more sustainable MP production under saline conditions using biogas, as an alternative to natural gas-based production with M. capsulatus . These strategic upstream choices could bring us closer to a future-proof protein production system.

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