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Key amino acid residues of the AGT 1 permease required for maltotriose consumption and fermentation by Saccharomyces cerevisiae
Author(s) -
Trichez D.,
Knychala M.M.,
Figueiredo C.M.,
Alves S.L.,
Silva M.A.,
Miletti L.C.,
Araujo P.S.,
Stambuk B.U.
Publication year - 2019
Publication title -
journal of applied microbiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.889
H-Index - 156
eISSN - 1365-2672
pISSN - 1364-5072
DOI - 10.1111/jam.14161
Subject(s) - maltotriose , permease , lactose permease , biochemistry , symporter , saccharomyces cerevisiae , mutant , yeast , amino acid , biology , chemistry , transporter , maltose , gene , enzyme
Aims The AGT 1 gene encodes for a general α‐glucoside‐H + symporter required for efficient maltotriose fermentation by Saccharomyces cerevisiae . In the present study, we analysed the involvement of four charged amino acid residues present in this transporter that are required for maltotriose consumption and fermentation by yeast cells. Methods and Results By using a knowledge‐driven approach based on charge, conservation, location, three‐dimensional (3D) structural modelling and molecular docking analysis, we identified four amino acid residues (Glu‐120, Asp‐123, Glu‐167 and Arg‐504) in the AGT 1 permease that could mediate substrate binding and translocation. Mutant permeases were generated by site‐directed mutagenesis of these charged residues, and expressed in a yeast strain lacking this permease ( agt1 ∆). While mutating the Arg‐504 or Glu‐120 residues into alanine totally abolished (R504A mutant) or greatly reduced (E120A mutant) maltotriose consumption by yeast cells, as well as impaired the active transport of several other α‐glucosides, in the case of the Asp‐123 and Glu‐167 amino acids, it was necessary to mutate both residues (D123G/E167A mutant) in order to impair maltotriose consumption and fermentation. Conclusions Based on the results obtained with mutant proteins, molecular docking and the localization of amino acid residues, we propose a transport mechanism for the AGT 1 permease. Significance and Impact of the Study Our results present new insights into the structural basis for active α‐glucoside‐H + symport activity by yeast transporters, providing the molecular bases for improving the catalytic properties of this type of sugar transporters.