Meta-Analysis and Functional Validation of Nutritional Requirements of Solventogenic Clostridia Growing under Butanol Stress Conditions and Coutilization ofd-Glucose andd-Xylose

Abstract

Recent advances in systems biology, omics, and computational studies allow us to carry out data mining for improving biofuel production bioprocesses. Of particular interest are bioprocesses that center on microbial capabilities to biotransform both the hexose and pentose fractions present in crop residues. This called for a systematic exploration of the components of the media to obtain higher-density cultures and more-productive fermentation operations than are currently found. By using a meta-analysis approach of the transcriptional responses to butanol stress, we identified the nutritional requirements of solvent-tolerant strain Clostridium beijerinckii SA-1 (ATCC 35702). The nutritional requirements identified were later validated using the chemostat pulse-and-shift technique. C. beijerinckii SA-1 was cultivated in a two-stage single-feed-stream continuous production system to test the proposed validated medium formulation, and the coutilization of D-glucose and D-xylose was evaluated by taking advantage of the well-known ability of solventogenic clostridia to utilize a large variety of carbon sources such as mono-, oligo-, and polysaccharides containing pentose and hexose sugars. Our results indicated that C. beijerinckii SA-1 was able to coferment hexose/pentose sugar mixtures in the absence of a glucose repression effect. In addition, our analysis suggests that the solvent and acid resistance mechanisms found in this strain are differentially regulated compared to strain NRRL B-527 and are outlined as the basis of the analysis toward optimizing butanol production.

Comparative phenotypic analysis and genome sequence of Clostridium beijerinckii SA-1, an offspring of NCIMB 8052

“…After activation, cultures were transferred into the same medium used in the reactors. The composition of the medium used for the batch experiments has been reported previously (Heluane et al, 2011). The medium base components were autoclaved without the sugars and trace components, which were always sterilized by filtration and added aseptically to the medium reservoir.…”

“…In SA-1, the SNV changed a histidine residue, positively charged, for a polar uncharged asparagine residue. This missense mutation at residue 47 (H47N) occurred in a conserved region of the protein and may have an effect on the function of the protein in regulating purine utilization (Xi et al, 2000;Heluane et al, 2011).…”

“…We compared the solventogenic production capabilities of NCIMB 8052 and SA-1 growing in a carbon-and energylimiting medium (Heluane et al, 2011), and further identified and confirmed genetic polymorphisms in their genome sequences.…”

“…This taxonomically complex group of organisms, when cultured under the right conditions, possesses a metabolic ability to convert a diverse group of carbohydrates, including pentoses, hexoses and even polysaccharides, into butanol and other subproducts (Mitchell, 1997;Qureshi et al, 2007Qureshi et al, , 2010; Lee et al, 2008;Jang et al, 2012a). Additionally, a large number of studies on ABE fermentation have been carried out demonstrating that among solventogenic strains, Clostridium beijerinckii exhibits unique cellulolytic and xylanolytic activities as well as desirable properties such as sugar cofermentation (Lee et al, 1985a, b;Wang & Blaschek, 2011;Heluane et al, 2011). These properties facilitate the exploitation by fermentation of alternative sustainable sugar-based feedstock that also contains vitamins and nitrogenous compounds required for successful ABE fermentations.…”

“…SA-1 has been overlooked the last three decades and only a few studies have evaluated this strain, including the work assessing its a-amylase and glucoamylase activities (Chojecki & Blaschek, 1986). Recently, we evaluated the nutritional requirements of SA-1 and proposed a medium that facilitates its growth under butanol stress, making it possible to obtain reliable continuous butanol production processes when the pH was kept at 6.5 (Heluane et al, 2011). Under these conditions SA-1 also exhibited characteristics of delayed lysis during sporulation similar to a Clostridium acetobutylicum lyt-l mutant -a desirable phenotype that improves butanol accumulation (Van Der Westhuizen et al, 1982).…”