O-029. Application of Systems Biology Tools to Unravel the Mechanisms of Acetate Tolerance in Zymomonas mobilis

S. Yang1, D. A. Pelletier1, C. Pan1, T. J. Tschaplinski1, S. L. Martin2, N. L. Engle1, G. B. Hurst1, N. F. Samatova1, Y. Yang1, S. D. Brown1;
1Oak Ridge Natl. Lab., Oak Ridge, TN, 2North Carolina State Univ., Raleigh, NC.

Zymomonas mobilis is a gram-negative ethanogenic bacterium that produces ethanol via the Entner-Doudoroff pathway. Z. mobilis is a good candidate for bioethanol production with desirable attributes of low biomass production, high sugar-uptake rates, and high ethanol yields and tolerance. Acetic acid has been identified as a major inhibitor present in lignocellulosic hydrolysates. A sodium acetate tolerant mutant strain (AcR) derived classically with described improvements in ethanol productivity in the presence of acetate was characterized using systems biology tools. The aim of these studies was to identify the molecular stress mechanisms and the targets for strain development. We compared the AcR and wild-type Z. mobilis ZM4 (ZM4) strains using microarray genome resequencing. Thirty-eight SNPs were identified including 26 found within coding regions and 12 within non-coding regions. In addition, 221 non-called region of interest (NCR) were also identified covering 16 regions with a one 1.4-kb deletion containing 206 NCR. PCR and traditional Sanger sequencing confirmed the deletion and SNPs. Transcriptomic profiles for AcR and ZM4 under acetate stress were generated and data suggested expression of one region was correlated with the observed genome structural changes. These data were further supported by a quantitative proteomics experiment comparing the AcR and ZM4 strains under acetate stress using 14/15N metabolic labeling. We identified 1,142 proteins out of possible 1,998 ZM4 proteins with several differentially expressed within different growth phases. The proteomics data also confirmed the identity of five mutant proteins in the AcR strain. In summary, we have identified loci related to the Z. mobilis acetate stress response, and are confirming their roles using the molecular and biochemistry approaches. We established a platform that integrates the advantages of classical selection strategies and systems biology technologies to establish a new paradigm in industrial strain characterization and development.