The ethanol-induced global alteration in Arthrobacter simplex and its mutants with enhanced ethanol tolerance

Arthrobacter simplex has received considerable interests due to its superior Δ 1 -dehydrogenation ability. Ethanol used as co-solvent is a stress commonly encountered during biotransformation. Therefore, studies of ethanol tolerance of A. simplex are of great importance to improve the biotransformation efficiency. In this paper, the combined analysis of physiological properties, cell compositions, stress-responsive metabolites, and proteome profiles was carried out to achieve a global view of ethanol tolerance of A. simplex. Under sublethal conditions, cell permeability and membrane fluidity exhibited concentration-dependent increase by affecting the contents or compositions of cell peptidoglycan, lipids, phospholipids, and fatty acids. Among them, cis – trans isomerization of unsaturated fatty acids was a short-term and reversible process, while the changes in phospholipid headgroups and increase in saturation degree of fatty acids were long-term and irreversible processes, which collectively counteracted the elevated membrane fluidity caused by ethanol and maintained the membrane stability. The decreased intracellular ATP content was observed at high ethanol concentration since proton motive force responsible for driving ATP synthesis was dissipated. The involvement of trehalose and glycerol, oxidative response, and DNA damage were implicated due to their changes in positive proportion to ethanol concentration. Proteomic data supported that ethanol invoked a global alteration, among which, the change patterns of proteins participated in the biosynthesis of cell wall and membrane, energy metabolism, compatible solute metabolism, and general stress response were consistent with observations from cell compositions and stress-responsive metabolites. The protective role of proteins participated in DNA repair and antioxidant system under ethanol stress was validated by overexpression of the related genes. This is the first demonstration on ethanol tolerance mechanism of A. simplex , and the current studies also provide targets to engineer ethanol tolerance of A. simplex .