Ribas David, Sá-Pessoa Joana, Soares-Silva Isabel, Paiva Sandra, Nygård Yvonne, Ruohonen Laura, Penttilä Merja, Casal Margarida
Centre of Molecular and Environmental Biology (CBMA), Department of Biology, University of Minho, Campus de Gualtar, Braga 4710-057, Portugal.
VTT Technical Research Centre of Finland Ltd, Tietotie 2, P.O. Box 1000, 02044 VTT Espoo, Finland.
FEMS Yeast Res. 2017 Mar 1;17(2). doi: 10.1093/femsyr/fox005.
Sugar acids can be used as platform chemicals to generate primary building blocks of industrially relevant products. Microbial production of these organic compounds at high yields requires the engineering of the enzymatic machinery and the presence of plasma membrane transporters able to export them outside the cells. In this study, several yeast carboxylic acid transporters belonging to the Jen family were screened for the transport of biotechnologically relevant sugar acids, namely gluconic, saccharic, mucic, xylaric and xylonic acid, and functionally characterised in Saccharomyces cerevisiae. We show that Jen permeases are capable of transporting most of these sugar acids, although with different specificities. Saccharate is a substrate of the transporters ScJen1-S271Q and KlJen2, gluconate of CaJen2 and KlJen2, and xylarate and mucate of CaJen2. A molecular docking approach of these transporters identified the residues that play a major role in the substrate binding of these sugar acids, namely R188 (ScJen1), R122 (CaJen2) and R127 (KlJen2), all equivalent residues (TMS II). The identification of Jen members as sugar acid transporters can contribute to engineering efficient microbial cell factories with increased sugar acid production, as the ScJen1 is able to promote substrate efflux.
糖酸可作为平台化学品，用于生成具有工业相关性产品的主要构建模块。要以高产率微生物生产这些有机化合物，需要对酶促机制进行工程改造，并存在能够将它们输出到细胞外的质膜转运蛋白。在本研究中，筛选了几种属于Jen家族的酵母羧酸转运蛋白，以检测其对具有生物技术相关性的糖酸（即葡萄糖酸、糖二酸、粘酸、木糖二酸和木糖酸）的转运能力，并在酿酒酵母中对其功能进行了表征。我们发现，Jen通透酶能够转运这些糖酸中的大多数，尽管具有不同的特异性。糖二酸是转运蛋白ScJen1-S271Q和KlJen2的底物，葡萄糖酸是CaJen2和KlJen2的底物，木糖二酸和粘酸是CaJen2的底物。对这些转运蛋白进行的分子对接分析确定了在这些糖酸底物结合中起主要作用的残基，即R188（ScJen1）、R122（CaJen2）和R127（KlJen2），均为等效残基（跨膜结构域II）。将Jen成员鉴定为糖酸转运蛋白有助于构建高效的微生物细胞工厂，提高糖酸产量，因为ScJen1能够促进底物外排。
