Metabolic Response of Clostridium ljungdahlii to Oxygen Exposure

Abstract

cClostridium ljungdahlii is an important synthesis gas-fermenting bacterium used in the biofuels industry, and a preliminary investigation showed that it has some tolerance to oxygen when cultured in rich mixotrophic medium. Batch cultures not only continue to grow and consume H 2 , CO, and fructose after 8% O 2 exposure, but fermentation product analysis revealed an increase in ethanol concentration and decreased acetate concentration compared to non-oxygen-exposed cultures. In this study, the mechanisms for higher ethanol production and oxygen/reactive oxygen species (ROS) detoxification were identified using a combination of fermentation, transcriptome sequencing (RNA-seq) differential expression, and enzyme activity analyses. The results indicate that the higher ethanol and lower acetate concentrations were due to the carboxylic acid reductase activity of a more highly expressed predicted aldehyde oxidoreductase (CLJU_c24130) and that C. ljungdahlii's primary defense upon oxygen exposure is a predicted rubrerythrin (CLJU_c39340). The metabolic responses of higher ethanol production and oxygen/ ROS detoxification were found to be linked by cofactor management and substrate and energy metabolism. This study contributes new insights into the physiology and metabolism of C. ljungdahlii and provides new genetic targets to generate C. ljungdahlii strains that produce more ethanol and are more tolerant to syngas contaminants. Clostridium ljungdahlii is an anaerobic, motile, endosporeforming, Gram-positive, rod-shaped, acetogenic bacterium isolated from chicken yard waste and has application as a biocatalyst to transform syngas components (CO, CO 2 , and H 2 ) into more valuable chemicals (1-4). It was the first bacterium discovered to metabolize these syngas components and to produce ethanol with acetate as its primary fermentation product (1, 4). To reduce the amount of acetate and increase the amount of ethanol produced by C. ljungdahlii for its use in industrial solvent production, different reactor designs and agitation parameters, varied syngas component concentrations, increased gas flow rates and pressures, reduced nitrogen sources, addition of reducing agents to media, adjustment of growth medium pH, and addition of nanoparticles have all been evaluated (1,(5)(6)(7)(8)(9)(10)(11)(12)(13)(14)(15). Some of these efforts have reportedly improved ethanol/ acetate product ratios from 1:20 to 2:1 in batch cultures and from 1:1 to 21:1 for cultures grown in continuously stirred reactors (8,9,16). More recent sequencing and genetic modification techniques have greatly enhanced our understanding of C. ljungdahlii's metabolism and increased production of ethanol as well as enabled production of other chemicals (e.g., acetone, butyrate, and butanol) (2, 3, 17-21).Despite these advancements, for C. ljungdahlii to be effectively used for industrial syngas transformation, some of its catalytic limitations related to gaseous headspace composition still require evaluation. Although steel mill waste gas was recently used as...

Biofilm Formation by Clostridium ljungdahlii Is Induced by Sodium Chloride Stress: Experimental Evaluation and Transcriptome Analysis

“…These results are in contrast to what was expected, since stress causes a slower metabolism, while ethanol production by C . ljungdahlii is often induced by stress from, for instance, a low pH or oxygen exposure [41, 57]. Our findings, therefore, might reflect different growth characteristics of the planktonic and the biofilm cells.…”

“…In addition, rubrerythrin, a non-haem Fe protein reported to be involved in the response of C . ljungdahlii to oxygen stress [41], was upregulated.…”

“…ljungdahlii [13], was strongly downregulated. Also a gene encoding a putative aldehyde oxidoreductase (CLJU_c24130), which was involved in the increased ethanol production after oxygen stress [41], was significantly downregulated (Table N in S2 File). These results are in contrast to what was expected, since stress causes a slower metabolism, while ethanol production by C .…”

Synthetic Biology on Acetogenic Bacteria for Highly Efficient Conversion of C1 Gases to Biochemicals

“…4). Through transcriptome analysis, it was possible to reveal the level of transcriptional expression of genes related to the WL pathway and the energy conservation system in acetogens under autotrophic conditions [ 36 , 105 , 106 , 107 , 108 , 109 , 110 ]. In addition, ribosome profiling in C. ljungdahlii or E. limosum revealed that translational regulation of genes in the carbonyl-branch or energy conservation system is critical to their autotrophic growth [ 111 , 112 ].…”

A high gas fraction, reduced power, syngas bioprocessing method demonstrated with a Clostridium ljungdahlii OTA1 paper biocomposite

“…Our model system uses Clostridium ljungdahlii OTA1 to convert CO from syngas as its sole carbon source into ethanol and acetate (Koepke et al, 2011; Ragsdale and Pierce, 2008; Tanner et al, 1993). Mutant OTA1 was derived from ATCC 55383 and isolated for its desirable ethanol to acetate ratio and CO consumption (Tirado-Acevedo, 2010; Whitham et al, 2015). Wild type C. ljungdahlii has been shown to have a maximum specific uptake of ~35 mmol CO g DCW −1 h −1 in liquid culture (Mohammadi et al, 2014; Richter et al, 2013).…”