Extracellular loops matter - subcellular location and function of the lysine transporter Lyp1 from Saccharomyces cerevisiae

FEBS J. 2020 Oct;287(20):4401-4414. doi: 10.1111/febs.15262. Epub 2020 Mar 11.

Abstract

Yeast amino acid transporters of the APC superfamily are responsible for the proton motive force-driven uptake of amino acids into the cell, which for most secondary transporters is a reversible process. The l-lysine proton symporter Lyp1 of Saccharomyces cerevisiae is special in that the Michaelis constant from out-to-in transport ( K m out in ) is much lower than K m in out , which allows accumulation of l-lysine to submolar concentration. It has been proposed that high intracellular lysine is part of the antioxidant mechanism of the cell. The molecular basis for the unique kinetic properties of Lyp1 is unknown. We compared the sequence of Lyp1 with APC para- and orthologues and find structural features that set Lyp1 apart, including differences in extracellular loop regions. We screened the extracellular loops by alanine mutagenesis and determined Lyp1 localization and activity and find positions that affect either the localization or activity of Lyp1. Half of the affected mutants are located in the extension of extracellular loop 3 or in a predicted α-helix in extracellular loop 4. Our data indicate that extracellular loops not only connect the transmembrane helices but also serve functionally important roles.

Keywords: APC superfamily; amino acid; loop; membrane transport; yeast.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Transport Systems, Basic / analysis
  • Amino Acid Transport Systems, Basic / genetics
  • Amino Acid Transport Systems, Basic / metabolism*
  • Computational Biology
  • Kinetics
  • Lysine / metabolism
  • Models, Molecular
  • Saccharomyces cerevisiae / chemistry
  • Saccharomyces cerevisiae / cytology
  • Saccharomyces cerevisiae / metabolism*
  • Saccharomyces cerevisiae Proteins / analysis
  • Saccharomyces cerevisiae Proteins / genetics
  • Saccharomyces cerevisiae Proteins / metabolism*

Substances

  • Amino Acid Transport Systems, Basic
  • LYP1 protein, S cerevisiae
  • Saccharomyces cerevisiae Proteins
  • Lysine