Influences of gaseous substrate composition on aerobic gas fermentation by Cupriavidus necator

Microbial gas fermentation is a promising biotechnology for the production of a variety of industrially relevant chemicals using ubiquitous gaseous feedstocks. Implementation of efficient fermentation processes requires an understanding of the microbial response to different ratios of the gaseous feedstocks (hydrogen, carbon dioxide, oxygen) supplied to the reactor. Here, we studied the effect of inlet gas composition on the characteristics of Cupriavidus necator , a common strain used in industrial biotechnology due to its metabolic diversity, genetic tractability, and ability to grow to high cell densities during aerobic fermentation. We investigated whether the supply of different proportions of hydrogen and carbon dioxide in the inlet gas stream (8:1, 3:1, and 1:1 H 2 :CO 2 ) while maintaining 13% dissolved oxygen affects the physiology of C. necator , with a focus on the accumulation of poly-3-hydroxybutyrate (PHB) and single-cell protein (SCP) within the microbial biomass. Biomass and PHB production were comparable between the 8:1 ratio (22 ± 5 g/L and 10.0 ± 0.4 g/L, respectively) and 3:1 ratio (18 ± 3 g/L and 5.8 ± 3.4 g/L, respectively) after seven days of incubation. Growth and PHB production were negatively impacted using the 1:1 ratio (8 ± 3 and 0.36 ± 1.08 g/L, respectively). SCP production varied as a function of both length of incubation and gas ratio supplied. We performed proteome profiling across time and treatment groups to assess relationships between productivity and protein expression. This study highlights the importance of gaseous substrate composition and its potential effects on microbial physiology and metabolism during fermentation. Key points • H 2 :CO 2 ratios of 8:1 and 3:1 produced comparable quantities of fermentation products. • Essential amino acids were enriched in SCP collected from the 1:1 fermentations. • Proteomics revealed differential expression in metabolic pathways of interest.