(491e) Zinc-Mediated Pleiotropic Effects and Regulatory Mechanism on Acetone-Butanol-Ethanol (ABE) Fermentation by Clostridium acetobutylicum

You-Duo Wu^1*, Chuang Xue¹ and Li-Jie Chen¹

¹School of Life Science and Biotechnology, Dalian University of Technology, Dalian, China

Recently, micronutrient zinc as a key cofactor has been demonstrated to play pleiotropic effects on cellular metabolism associated with differential carbohydrate utilization, cell growth, stress response as well as metabolic transition of acidogenesis and solventogenesis. Therefore, uncovering the zinc-mediated regulatory mechanism on preferred cellular traits was essential for exploring beneficial interactions of responsive genes and expanding regulatory network at system levels. The potential regulatory network involving in multigenic interactions and multiple cellular subsystems will contribute to the sustainable improvement of genetic and metabolic engineered solventogenic clostridia.

LC-ESI/MS analysis for intracellular metabolites revealed that, despite of rapid glucose utilization, glycolysis-derived metabolites such as G6P, F6P, FBP and PEP were dramatically decreased along with increased levels of energy supply ATP and reducing power NADH at the early stage of glucose-based ABE fermentation, which could well account for the enhanced exponential growth and earlier initiation of solventogenesis due to excessive NADH-derived oxido-reduction re-balance. In addition, branch node intermediates such as AcCoA, AcAcCoA and BuCoA involved in carbon flux distribution of acidogenesis/solventogenesis were also decreased coupled with extracellular products re-distribution, suggesting that micronutrient zinc could facilitate glycolysis and central carbon metabolism.

Global transcriptional analysis demonstrated that the expression levels of mutiple genes responsible for carbohydrate transport/metabolism were differentially upregulated with zinc supplementation, including glucose-PTS genes glcG, CAC1353 and CAC1354, fructose operon fru and 14 genes involved in xylose and arabinose transport/metabolism. Among the glycolytic genes, glcK, pfkA, pgK and pykA expression levels were slightly upregulated while H₂ biosynthetic gene hydA downregulated, which was consistent with the enhanced glycolysis and inhibited hydrogenesis. Furthermore, several key acidogenic and solventogenic genes thlA, etfAB, bcd, crt, ctfAB, adc and bdhB were differentially upregulated, which accelerate cascading conversion of central carbon metabolism, indicating the zinc-mediated regulatory mechanism involved in metabolic and genetic network of C. acetobutylicum at global levels.

Key Words: Clostridium acetobutylicum; Micronutrient zinc; ABE fermentation; Intracellular metabolites analysis; Transcriptional analysis.