Abstract
BackgroundAcetic acid is a predominant by-product of lignocellulosic biofuel process, which inhibits microbial biocatalysts. Development of bacterial strains that are tolerant to acetic acid is challenging due to poor understanding of the underlying molecular mechanisms.ResultsIn this study, we generated and characterized two acetic acid-tolerant strains of Zymomonas mobilis using N-methyl-N′-nitro-N-nitrosoguanidine (NTG)-acetate adaptive breeding. Two mutants, ZMA-142 and ZMA-167, were obtained, showing a significant growth rate at a concentration of 244 mM sodium acetate, while the growth of Z. mobilis ATCC 31823 were completely inhibited in presence of 195 mM sodium acetate. Our data showed that acetate-tolerance of ZMA-167 was attributed to a co-transcription of nhaA from ZMO0117, whereas the co-transcription was absent in ATCC 31823 and ZMA-142. Moreover, ZMA-142 and ZMA-167 exhibited a converstion rate (practical ethanol yield to theorical ethanol yield) of 90.16% and 86% at 195 mM acetate-pH 5 stress condition, respectively. We showed that acid adaptation of ZMA-142 and ZMA-167 to 146 mM acetate increased ZMA-142 and ZMA-167 resulted in an increase in ethanol yield by 32.21% and 21.16% under 195 mM acetate-pH 5 stress condition, respectively.ConclusionThe results indicate the acetate-adaptive seed culture of acetate-tolerant strains, ZMA-142 and ZMA-167, could enhance the ethanol production during fermentation.
Highlights
Acetic acid is a predominant by-product of lignocellulosic biofuel process, which inhibits microbial biocatalysts
ZM481 was maintained in Rich medium (RM; 20.0 Glucose utilization (g/L) glucose, 10.0 g/L yeast Extract, 2.0 g/L KH2PO4, pH 5.0) at 30 °C without shaking
Seed cultures were grown at 30 °C without shaking to mid growth phase and subsequently inoculated at 10%(v/v) into fermentation medium (FM), which was identical in composition to the seed medium except that final concentrations of up to 196 mM sodium acetate (Ajax Chemicals, NSW, Australia, AR grade)
Summary
Acetic acid is a predominant by-product of lignocellulosic biofuel process, which inhibits microbial biocatalysts. Development of bacterial strains that are tolerant to acetic acid is challenging due to poor understanding of the underlying molecular mechanisms. The main challenge is suggested to be pretreatment and enzymatic hydrolysis steps with which fermentable sugars are released from the biomass. These steps have been suggested to generate a variety of toxic compounds and form a stressful environment which inhibits microbial fermentations [2,3,4]. Strategies have been developed to overcome such issues for high ethanol production including removal of fermentation inhibitors and use of inhibitor-tolerant strains. There is currently no ideal bacterial ethanologen available for use in an industrial setting
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