Evolution and functional diversification of yeast sugar transporters.

While simple sugars such as monosaccharides and disaccharide are the typical carbon source for most yeasts, whether a species can grow on a particular sugar is generally a consequence of presence or absence of a suitable transporter to enable its uptake. The most common transporters that mediate sugar import in yeasts belong to the major facilitator superfamily (MFS). Some of these, for example the Saccharomyces cerevisiae Hxt proteins have been extensively studied, but detailed information on many others is sparce. In part, this is because there are many lineages of MFS transporters that are either absent from, or poorly represented in, the model S. cerevisiae, which actually has quite a restricted substrate range. It is important to address this knowledge gap to gain better understanding of the evolution of yeasts and to take advantage of sugar transporters to exploit or engineer yeasts for biotechnological applications. This article examines the full repertoire of MFS proteins in representative budding yeasts (Saccharomycotina). A comprehensive analysis of 139 putative sugar transporters retrieved from 10 complete genomes sheds new light on the diversity and evolution of this family. Using the phylogenetic lens, it is apparent that proteins have often been misassigned putative functions and this can now be corrected. It is also often seen that patterns of expansion of particular genes reflects the differential importance of transport of specific sugars (and related molecules) in different yeasts, and this knowledge also provides an improved resource for the selection or design of tailored transporters.

Identification of novel pentose transporters in Kluyveromyces marxianus using a new screening platform.

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.

Hexose transport in Torulaspora delbrueckii: identification of Igt1, a new dual-affinity transporter.

Torulaspora delbrueckii is a yeast species receiving increasing attention from the biotechnology industry, with particular relevance in the wine, beer and baking sectors. However, little is known about its sugar transporters and sugar transport capacity, frequently a rate-limiting step of sugar metabolism and efficient fermentation. Actually, only one glucose transporter, Lgt1, has been characterized so far. Here we report the identification and characterization of a second glucose transporter gene, IGT1, located in a cluster, upstream of LGT1 and downstream of two other putative hexose transporters. Functional characterization of IGT1 in a Saccharomyces cerevisiae hxt-null strain revealed that it encodes a transporter able to mediate uptake of glucose, fructose and mannose and established that its affinity, as measured by Km, could be modulated by glucose concentration in the medium. In fact, IGT1-transformed S. cerevisiae hxt-null cells, grown in 0.1% glucose displayed biphasic glucose uptake kinetics with an intermediate- (Km = 6.5 ± 2.0 mM) and a high-affinity (Km = 0.10 ± 0.01 mM) component, whereas cells grown in 2% glucose displayed monophasic kinetics with an intermediate-affinity (Km of 11.5 ± 1.5 mM). This work contributes to a better characterization of glucose transport in T. delbrueckii, with relevant implications for its exploitation in the food industry.