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Characterization of AtNST‐KT1, a novel UDP‐galactose transporter from Arabidopsis thaliana
Author(s) -
Rollwitz Inga,
Santaella Marcella,
Hille Diana,
Flügge Ulf-Ingo,
Fischer Karsten
Publication year - 2006
Publication title -
febs letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.593
H-Index - 257
eISSN - 1873-3468
pISSN - 0014-5793
DOI - 10.1016/j.febslet.2006.06.082
Subject(s) - nucleotide sugar , biochemistry , nucleotide , complementary dna , biology , affinity chromatography , yeast , sugar phosphates , golgi apparatus , fructose , saccharomyces cerevisiae , endoplasmic reticulum , chemistry , enzyme , gene
Nucleotide sugar transporters (NST) mediate the transfer of nucleotide sugars from the cytosol into the lumen of the endoplasmatic reticulum and the Golgi apparatus. Because the NSTs show similarities with the plastidic phosphate translocators (pPTs), these proteins were grouped into the TPT/NST superfamily. In this study, a member of the NST‐KT family, AtNST‐KT1, was functionally characterized by expression of the corresponding cDNA in yeast cells and subsequent transport experiments. The histidine‐tagged protein was purified by affinity chromatography and reconstituted into proteoliposomes. The substrate specificity of AtNST‐KT1 was determined by measuring the import of radiolabelled nucleotide mono phosphates into liposomes preloaded with various unlabelled nucleotide sugars. This approach has the advantage that only one substrate has to be used in a radioactively labelled form while all the nucleotide sugars can be provided unlabelled. It turned out that AtNST‐KT1 represents a monospecific NST transporting UMP in counterexchange with UDP‐Gal but did not transport other nucleotide sugars. The AtNST‐KT1 gene is ubiquitously expressed in all tissues. AtNST‐KT1 is localized to Golgi membranes. Thus, AtNST‐KT1 is most probably involved in the synthesis of galactose‐containing glyco‐conjugates in plants.