School of Microbiology, Environmental Research Institute, APC Microbiome Institute, University College Cork, Cork T12 K8AF, Ireland.
Centre of Environmental and Molecular Biology, Department of Biology, University of Minho, Campus of Gualtar, R. da Universidade, Braga 4710-057, Portugal.
FEMS Yeast Res. 2021 May 10;21(4). doi: 10.1093/femsyr/foab026.
The capacity of yeasts to assimilate xylose or arabinose is strongly dependent on plasma membrane transport proteins. Because pentoses comprise a substantial proportion of available sugars in lignocellulosic hydrolysates, their utilisation is centrally important for the development of second generation biorefineries. Relatively few native pentose transporters have been studied and there is intense interest in expanding the repertoire. To aid the identification of novel transporters, we developed a screening platform in the native pentose-utilising yeast Kluyveromyces marxianus. This involved the targeted deletion of twelve transporters of the major facilitator superfamily (MFS) and application of a synthetic biology pipeline for rapid testing of candidate pentose transporters. Using this K. marxianus ΔPT platform, we identified several K. marxianus putative xylose or arabinose transporter proteins that recovered a null strain's ability to growth on these pentoses. Four proteins of the HGT-family were able to support growth in media with high or low concentrations of either xylose or arabinose, while six HXT-like proteins displayed growth only at high xylose concentrations, indicating solely low affinity transport activity. The study offers new insights into the evolution of sugar transporters in yeast and expands the set of native pentose transporters for future functional and biotechnological studies.
酵母同化木糖或阿拉伯糖的能力强烈依赖于质膜转运蛋白。因为戊糖在木质纤维素水解物中占可用糖的很大一部分，所以它们的利用对于第二代生物精炼厂的发展至关重要。相对较少的天然戊糖转运蛋白已经被研究过，并且人们对扩大其种类有着浓厚的兴趣。为了帮助鉴定新的转运蛋白，我们在天然戊糖利用酵母克鲁维酵母中开发了一个筛选平台。这涉及到靶向删除十二种主要易化因子超家族（MFS）的转运蛋白，并应用合成生物学管道快速测试候选戊糖转运蛋白。使用这个 K. marxianus ΔPT 平台，我们鉴定了几种 K. marxianus 假定的木糖或阿拉伯糖转运蛋白，它们恢复了缺失菌株在这些戊糖上生长的能力。四种 HGT 家族的蛋白能够在高或低浓度的木糖或阿拉伯糖培养基中支持生长，而六种 HXT 样蛋白仅在高木糖浓度下显示生长，表明仅具有低亲和力的转运活性。该研究为酵母中糖转运蛋白的进化提供了新的见解，并为未来的功能和生物技术研究扩展了天然戊糖转运蛋白的集合。
