Functional expression of the human UDP-galactose transporters in the yeast Saccharomyces cerevisiae

We describe the functional expression of the putative human Golgi UDP-galactose transporters (hUGT1 and hUGT2) in the yeast Saccharomyces cerevisiae, Both hUGT1 and hUGT2 were expressed under the control of the yeast constitutive GAPDH promoter. The expression level of hUGT1 seemed tee be considerably lower than that of hUGT2, although hUGT1 has an amino acid sequence identical to that of hUGT2 except for 5 amino acid residues at the C-terminus. The hUGT product was expressed in the membranes of Golgi and other organellar compartments. The membrane vesicles prepared from the hUGT1- or the hUGT2-expressing yeast cells exhibited UDP-galactose specific transport activity. The apparent K-m values of the yeast-expressed hUGT1 and hUGT2 for UDP-galactose were 1.2 and 2 mu M, respectively, which were comparable with the K-m obtained with mammalian Golgi vesicles. Transport was dependent on temperature and integrity of vesicles, sm(a was inhibited by UMP, as observed with mammalian Golgi vesicles. Our results demonstrate that the previously described hUGT1 and hUGT2 encode the UDP-galactose transporters, rather than regulatory proteins, The development of a convenient yeast expression system Should facilitate analysis of the structure-function relationships of the UDP-galactose transporters.