CD-Laboratory for Biotechnology of Glycerol, University of Natural Resources and Life Sciences Vienna, Muthgasse 18, 1190 Vienna, Austria.
Institute of Microbiology and Microbial Biotechnology, Department of Biotechnology, BOKU-University of Natural Resources and Life Sciences Vienna, Muthgasse 18, 1190 Vienna, Austria.
FEMS Yeast Res. 2020 Oct 15;20(7). doi: 10.1093/femsyr/foaa055.
Yarrowia lipolytica is a yeast with many talents, one of them being the production of citric acid. Although the citrate biosynthesis is well studied, little is known about the transport mechanism by which citrate is exported. To gain better insight into this mechanism, we set out to identify a transporter involved in citrate export of Y. lipolytica. A total of five proteins were selected for analysis based on their similarity to a known citrate exporter, but neither a citrate transport activity nor any other phenotypic function could be attributed to them. Differential gene expression analysis of two strains with a distinct citrate productivity revealed another three putative transporters, one of which is YALI0D20196p. Disrupting YALI0D20196g in Y. lipolytica abolished citrate production, while extrachromosomal expression enhanced citrate production 5.2-fold in a low producing wildtype. Furthermore, heterologous expression of YALI0D20196p in the non-citrate secreting yeast Saccharomyces cerevisiae facilitated citrate export. Likewise, expression of YALI0D20196p complemented the ability to secrete citrate in an export-deficient strain of Aspergillus niger, confirming a citrate export function of YALI0D20196p. This report on the identification of the first citrate exporter in Y. lipolytica, termed Cex1, represents a valuable starting point for further investigations of the complex transport processes in yeasts.
解脂耶氏酵母是一种多才多艺的酵母，其中之一是生产柠檬酸。尽管柠檬酸的生物合成已经得到了很好的研究，但对于柠檬酸的输出运输机制知之甚少。为了更好地了解这一机制，我们着手鉴定一种参与解脂耶氏酵母柠檬酸输出的转运蛋白。总共选择了 5 种基于其与已知柠檬酸外排蛋白相似性的蛋白质进行分析，但它们既没有柠檬酸转运活性，也没有其他表型功能。对两种具有不同柠檬酸生产力的菌株的差异基因表达分析揭示了另外三个假定的转运蛋白，其中一个是 YALI0D20196p。在解脂耶氏酵母中敲除 YALI0D20196g 会导致柠檬酸生产完全丧失，而在外源染色体上表达则可使低产野生型的柠檬酸产量提高 5.2 倍。此外，在非柠檬酸分泌酵母酿酒酵母中异源表达 YALI0D20196p 有助于柠檬酸的外排。同样，在柠檬酸分泌缺陷型黑曲霉菌株中表达 YALI0D20196p 可补充其分泌柠檬酸的能力，证实了 YALI0D20196p 具有柠檬酸外排功能。本报告鉴定了解脂耶氏酵母中的第一个柠檬酸外排蛋白 Cex1，这为进一步研究酵母中复杂的转运过程提供了有价值的起点。
