A substrate translocation trajectory in a cytoplasm-facing topological model of the monocarboxylate/H+ symporter Jen1p

Isabel Soares-Silva, Joana Sá-Pessoa, Vassilios Myrianthopoulos, Emmanuel Mikros, Margarida Casal*, George Diallinas

*Corresponding author for this work

Research output: Contribution to journalArticle

17 Citations (Scopus)

Abstract

Previous mutational analysis of Jen1p, a Saccharomyces cerevisiae monocarboxylate/H+ symporter of the Major Facilitator Superfamily, has suggested that the consensus sequence 379NXX[S/T]HX[S/T]QD387 in transmembrane segment VII (TMS-VII) is part of the substrate translocation pathway. Here, we rationally design, analyse and show that several novel mutations in TMS-V and TMS-XI directly modify Jen1p function. Among the residues studied, F270 (TMS-V) and Q498 (TMS-XI) are critical specificity determinants for the distinction of mono- from dicarboxylates, and N501 (TMS-XI) is a critical residue for function. Using a model created on the basis of Jen1p similarity with the GlpT permease, we show that all polar residues critical for function within TMS-VII and TMS-XI (N379, H383, D387, Q498, N501) are perfectly aligned in an imaginary axis that lies parallel to the protein pore. This model and subsequent mutational analysis further reveal that an additional polar residue facing the pore, R188 (TMS-II), is irreplaceable for function. Our model also justifies the role of F270 and Q498 in substrate specificity. Finally, docking calculations reveal a 'trajectory-like' substrate displacement within the Jen1p pore, where R188 plays a major dynamic role mediating the orderly relocation of the substrate by subsequent H-bond interactions involving itself and residues H383, N501 and Q498.

Original languageEnglish
Pages (from-to)805-817
Number of pages13
JournalMolecular Microbiology
Volume81
Issue number3
DOIs
Publication statusPublished - 01 Aug 2011
Externally publishedYes

ASJC Scopus subject areas

  • Microbiology
  • Molecular Biology

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