Project description:Kluyveromyces marxianus is a non-conventional yeast with outstanding physiological characteristics and a high potential for lignocellulosic ethanol production. However, achieving high ethanol produc-tivities implies overcoming several biotechnological challenges as the cellular inhibition caused by the inhibitors present in the medium. In this work, the adaptation of K. marxianus SLP1 to increase the tolerance to a mix of inhibitory compounds was carried out using the adaptive laboratory evolution (ALE) strategy. As a result of the ALE adaptation process, an improved K. marxianus strain (P8) was obtained after 8 serial passes. The fermentative and physiological parameters evidenced a better response of the P8 strain against the synergistic effect of multiple inhibitors. The P8 strain reduced the lag phase from 12 to 4 h, increasing 40% the biomass and improving 16-fold the volumetric eth-anol productivity. To test the transcriptional dynamics for the adaptation process, we performed a differential gene expression analysis in control conditions; the results showed that the basal gene expression in P8 changes, suggesting the biological capability of K. marxianus to activate the ad-aptative prediction mechanism. This study demonstrates the rapid adaptability of K. marxianus SLP1 to stressful environments, making this yeast a promising candidate to produce lignocellulosic ethanol.
Project description:Organisms specialized to extreme environments can be the product of millions of years of evolutionary engineering and refinement. The underlying genetics can be quite distinct from those operating at earlier stages of trait innovation. In this work, we have developed the multistress-resistant yeast Kluyveromyces marxianus, which diverged from its closest relative >20 million years ago, as a model for interspecies comparative biology and genomics. In growth assays of the Kluyveromyces genus, we found that K. marxianus exhibited unique tolerance of high heat and a subset of chemical stress conditions. We then generated and analyzed omic profiles from across the genus to find molecular features associated with—and potentially causal for—K. marxianus traits. Expression profiling revealed divergent lipid processing and membrane transport programs in K. marxianus, borne out in changes in lipid utilization in experimental assays. Sequence analyses found robust evidence for expansions in gene families in the K. marxianus genome, most notably among transmembrane transporters and in metabolic enzymes. In molecular-evolution tests, we identified adaptive protein variants throughout the K. marxianus genome, among which plasma membrane transporters were over-represented. These data enable a model of the molecular mechanisms and evolutionary pressures underlying K. marxianus traits, including adaptive changes to transporters, lipid processing, and membrane functions mediating stress resistance.
Project description:Thermotolerant Kluyveromyces marxianus possesses intrinsic abilities to ferment and assimilate a wide variety of substrates including xylose and to efficiently produce proteins. The transcriptome analysis clarified distinctive metabolic pathways under three different growth conditions, static culture, high temperature and xylose medium, in comparison to the control condition of glucose medium under a shaking condition at 30°C. Interestingly, the yeast appears to overcome the issue of reactive oxygen species, which tend to accumulate under all three conditions.