Amino acid residues involved in ligand preference of the Snf3 transporter-like sensor in Saccharomyces cerevisiae.
Snf3 is a plasma membrane protein in Saccharomyces cerevisiae able to sense the presence of glucose. Although the Snf3 protein does not transport sugars, it shares sequence similarity with various glucose transporters from other organisms. We investigated the sugar specificity/preferences of Snf3. The ability of cells to sense sugars in vivo was monitored by following the degradation of the Mth1 protein, an early event in the signal pathway. Our study reveals that Snf3, in addition to glucose, also senses fructose and mannose, as well as the glucose analogues 2-deoxyglucose, 3-O-methylglucoside and 6-deoxyglucose. The signalling proficiency of a non-phosphorylatable analogue strongly supports the notion that sensing through Snf3 does not require sugar phosphorylation. Sequence comparisons of Snf3 to glucose transporters indicated amino acid residues possibly involved in sensing of sugars other than glucose. By site-specific mutagenesis of the structural gene, roles of specific residues in Snf3 could be established. Change of isoleucine-374 to valine in transmembrane segment 7 of Snf3 partially abolished sensing of fructose and mannose, while mutagenesis causing a change of phenylalanine-462 to tyrosine in transmembrane segment 10 of Snf3 abolished sensing of fructose. Neither of these amino acid changes affected the ability of Snf3 to sense glucose, nor did they permit Snf3 to sense galactose. These data indicate a similarity between a ligand binding site of the sensor Snf3 and binding sites used for facilitated hexose transport in the GLUT proteins.
Dietvorst
Judith
J
Carlsberg Laboratory, Gamle Carlsberg Vej 10, DK-2500 Copenhagen Valby, Denmark.
Karhumaa
Kaisa
K
Kielland-Brandt
Morten C
MC
Brandt
Anders
A
D048868:Adaptor Proteins, Signal Transducing / Q000378:metabolism; D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D000818:Animals; D050260:Carbohydrate Metabolism; D008024:Ligands; D009004:Monosaccharide Transport Proteins / Q000235:genetics* / Q000378:metabolism*; D016297:Mutagenesis, Site-Directed; D050505:Mutant Proteins / Q000235:genetics / Q000378:metabolism; D011485:Protein Binding; D012441:Saccharomyces cerevisiae / Q000235:genetics* / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics* / Q000378:metabolism*; D017386:Sequence Homology, Amino Acid; D013379:Substrate Specificity
D048868:Adaptor Proteins, Signal Transducing; D008024:Ligands; C118806:MTH1 protein, S cerevisiae; D009004:Monosaccharide Transport Proteins; D050505:Mutant Proteins; C104463:SNF3 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins