Enhanced Benzaldehyde Tolerance in Zymomonas mobilis Biofilms and the Potential of Biofilm Applications in Fine-Chemical Production

Abstract

Biotransformation plays an increasingly important role in the industrial production of fine chemicals due to its high product specificity and low energy requirement. One challenge in biotransformation is the toxicity of substrates and/or products to biocatalytic microorganisms and enzymes. Biofilms are known for their enhanced tolerance of hostile environments compared to planktonic free-living cells. Zymomonas mobilis was used in this study as a model organism to examine the potential of surface-associated biofilms for biotransformation of chemicals into value-added products. Z. mobilis formed a biofilm with a complex three-dimensional architecture comprised of microcolonies with an average thickness of 20 m, interspersed with water channels. Microscopic analysis and metabolic activity studies revealed that Z. mobilis biofilm cells were more tolerant to the toxic substrate benzaldehyde than planktonic cells were. When exposed to 50 mM benzaldehyde for 1 h, biofilm cells exhibited an average of 45% residual metabolic activity, while planktonic cells were completely inactivated. Three hours of exposure to 30 mM benzaldehyde resulted in sixfold-higher residual metabolic activity in biofilm cells than in planktonic cells. Cells inactivated by benzaldehyde were evenly distributed throughout the biofilm, indicating that the resistance mechanism was different from mass transfer limitation. We also found that enhanced tolerance to benzaldehyde was not due to the conversion of benzaldehyde into less toxic compounds. In the presence of glucose, Z. mobilis biofilms in continuous cultures transformed 10 mM benzaldehyde into benzyl alcohol at a steady rate of 8.11 g (g dry weight)؊1 day ؊1 with a 90% molar yield over a 45-h production period.Microbes in nature are commonly in surface-associated, matrix-enclosed structures referred to as biofilms (7,28). Bacteria in biofilms can differ profoundly in terms of phenotypic characteristics (31, 33) or adaptive responses to stress (17, 38) from their planktonic counterparts, which can provide survival advantages and protection in a range of environmental conditions (16,23). Biofilms are also commonly found on medical implants and are responsible for many persistent infections due to their increased resistance to antimicrobial agents (13).While deleterious in clinical settings, biofilms are of great interest in biotechnology. For example, biofilms have been used as biocatalysts in bioremediation and wastewater treatment processes (21,23,27) and in fermentative vinegar (9) and ethanol (20) production from agricultural materials, as well as in biosynthesis of polymers (42). To our knowledge, the use of biofilms for transformation of fine chemicals into value-added products has not been reported previously.Biotransformation using viable cells capable of cofactor regeneration is an important approach to fine-chemical production (30, 34). A common obstacle in biocatalytic fine-chemical production is the potential toxic effects of substrates and/or products on cells during the process. S...

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