Biology of Pentose-Fermenting Yeasts

在水解时，来自植物生物质的木质纤维素材料产生己糖（葡萄糖、甘露糖、半乳糖）和戊糖（木糖、阿拉伯糖）糖的混合物。 其中，葡萄糖是最丰富的，其次是木糖，而其他糖以低得多的浓度存在。 为了提高生物质转化过程的经济性，潜在的生物转化微生物必须能够有效地将己糖和戊糖发酵成燃料或化学品。 不幸的是，众所周知的和充分研究的发酵酵母酿酒酵母（Saccharomyces cerevisiae）不能利用木糖作为生长的碳源，尽管它具有木糖代谢所需的所有酶。 20世纪80年代初，加拿大和美国研究人员发现了戊糖发酵酵母，这是一个重要的里程碑，因为它给了人们戊糖可以发酵的希望。 然而，对戊糖发酵酵母的研究发现，在预处理衍生的抑制剂存在下，它们遭受葡萄糖抑制、低乙醇耐受性和较差的性能。我已经研究戊糖发酵酵母32年了，我仍然对我能从这些迷人的微生物中学到的一些新东西感到惊讶。 我的研究一直集中在这些酵母的生物化学，生理学和遗传改良上，这仍然是我的长期目标。 在本提案中，我计划研究与这些酵母相关的两个基本问题。 第一个与某些戊糖发酵酵母的低抑制剂耐受性有关。 本研究拟采用经典的随机诱变方法，提高木糖醇产生菌季也蒙假丝酵母FTI 20037对木质纤维素水解产物中抑制剂的耐受性。 将对改良的突变体进行全基因组测序分析，以鉴定共同赋予抑制剂耐受性的所有突变。 第二个问题与为什么一些戊糖发酵酵母表现出如此低的乙醇耐受性有关。 许多戊糖发酵酵母只能产生高达5%（w/v）的乙醇，而一些葡萄酒酵母可以产生高达10%（w/v）的乙醇。 我计划研究质膜结合酶ATP酶是否在乙醇耐受性中起作用，从而确定不同酵母可以产生的最大乙醇量。 我打算雇两个研究生来帮我完成这些有趣的项目。学生将接受酵母生理学，发酵以及生物质转化的培训。 这些基础研究的结果将提供有关这些有趣的微生物的有价值的知识，这些信息最终可能有助于制定改善戊糖发酵的策略。 研究结果还应与对酒精毒性和耐受性感兴趣的各方有关。

On hydrolysis, lignocellulosic materials from plant biomass yield a mixture of hexose (glucose, mannose, galactose) and pentose (xylose, arabinose) sugars. Among these, glucose is the most abundant, followed by xylose, while the other sugars are present in much lower concentrations. To improve the economics of a biomass conversion process, it is imperative that potential bioconversion microorganisms be able to ferment both the hexoses and pentoses efficiently into fuels or chemicals. Unfortunately, the well-known and well-studied fermentative yeast Saccharomyces cerevisiae is unable to utilize xylose as a carbon source for growth, despite the fact that it possesses all the enzymes needed for xylose metabolism. The discovery of pentose-fermenting yeasts in the early 1980s by Canadian and US researchers was a significant milestone as it gave people the hope that pentoses can be fermented. However, studies with pentose-fermenting yeasts found that they suffer from glucose repression, low ethanol tolerance and poor performance in the presence of pretreatment-derived inhibitors. I have been studying pentose-fermenting yeasts for 32 years and I continue to be amazed at some of the new things I can learn from these fascinating microorganisms. My research has been focused on the biochemistry, physiology and genetic improvement of these yeasts, and this continues to be my long term goal. In this proposal, I plan to study two fundamental issues related to these yeasts. The first is related to the low inhibitor tolerance of some pentose-fermenting yeasts. I plan to use classical random mutagenesis method to improve the tolerance of the xylitol-producing Candida guilliermondii FTI 20037 to inhibitors found in lignocellulosic hydrolysates. Improved mutants will be subjected to whole genome sequencing analysis to identify all the mutations that collectively confer inhibitor tolerance. The second is related to the question as to why some pentose-fermenting yeasts exhibit such low ethanol tolerance. Many pentose-fermenting yeasts can produce up to 5% (w/v) ethanol only while some wine yeasts can produce up to 10% (w/v) ethanol. I plan to investigate if the plasma membrane bound enzyme ATPase may play a role in ethanol tolerance and hence in determining the maximum amount of ethanol that can be produced by different yeasts. I plan to hire two graduate students to help me tackle these fun projects. The students will be trained in yeast physiology, fermentation as well as biomass conversion. Results forthcoming from these fundamental studies will provide valuable knowledge about these interesting microorganisms and the information may ultimately be useful to devise strategies to improve pentose fermentation. The results should also be relevant to parties interested in alcohol toxicity and tolerance in general.