Quantitative proteomic analysis reveals the ethanologenic metabolism regulation of by exogenous ethanol addition.

Background: H-ethanol-coproducing bacteria, as primary fermenters, play important roles in the microbiome of bioreactors for bioenergy production from organic wastewater or solid wastes. YUAN-3 is an anaerobic ethanol-H-fermenting bacterium. Ethanol is one of the main end-products of strain YUAN-3 that influence its fermentative process. Until recently, the molecular mechanism of metabolic regulation in strain YUAN-3 during ethanol accumulation has still been unclear. This study aims to elucidate the metabolic regulation mechanisms in strain YUAN-3, which contributes to effectively shape the microbiome for biofuel and bioenergy production from waste stream.

Results: This study reports that ethanol stress altered the distribution of end-product yields in the H-ethanol-coproducing strain YUAN-3. Decreasing trends of hydrogen yield from 1888.6 ± 45.8 to 837 ± 64.7 mL L and acetic acid yield from 1767.7 ± 45 to 160.6 ± 44.7 mg L were observed in strain YUAN-3 with increasing exogenous ethanol (0 mM-200 mM). However, the ethanol yield of strain YUAN-3 increased by 15.1%, 30.1%, and 27.4% in 50 mM, 100 mM, and 200 mM ethanol stress, respectively. The endogenous ethanol accounted for 96.1% (w/w) in liquid end-products when exogenous ethanol of 200 mM was added. The molar ratio of ethanol to acetic acid increased 14 times (exogenous ethanol of 200 mM) compared to the control. iTRAQ-based quantitative proteomic analysis indicated that 263 proteins of strain YUAN-3 were differentially expressed in 50 mM, 100 mM, and 200 mM of exogenous ethanol. These proteins are mainly involved in amino acid transport and metabolism, central carbon metabolism, and oxidative stress response.

Conclusion: These differentially expressed proteins play important roles in metabolic changes necessary for growth and survival of strain YUAN-3 during ethanol stress. The up-regulation of bifunctional acetaldehyde-CoA/alcohol dehydrogenase (ADHE) was the main reason why ethanol production was enhanced, while hydrogen gas and acetic acid yields declined in strain YUAN-3 during ethanol stress. This study also provides a new approach for the enhancement of ethanologenesis by H-ethanol-coproducing bacteria through exogenous ethanol addition.