Structural bases for the interaction and stabilization of the human amino acid transporter LAT2 with its ancillary protein 4F2hc

Proc Natl Acad Sci U S A. 2014 Feb 25;111(8):2966-71. doi: 10.1073/pnas.1323779111. Epub 2014 Feb 10.

Abstract

Heteromeric amino acid transporters (HATs) are the unique example, known in all kingdoms of life, of solute transporters composed of two subunits linked by a conserved disulfide bridge. In metazoans, the heavy subunit is responsible for the trafficking of the heterodimer to the plasma membrane, and the light subunit is the transporter. HATs are involved in human pathologies such as amino acidurias, tumor growth and invasion, viral infection and cocaine addiction. However structural information about interactions between the heavy and light subunits of HATs is scarce. In this work, transmission electron microscopy and single-particle analysis of purified human 4F2hc/L-type amino acid transporter 2 (LAT2) heterodimers overexpressed in the yeast Pichia pastoris, together with docking analysis and crosslinking experiments, reveal that the extracellular domain of 4F2hc interacts with LAT2, almost completely covering the extracellular face of the transporter. 4F2hc increases the stability of the light subunit LAT2 in detergent-solubilized Pichia membranes, allowing functional reconstitution of the heterodimer into proteoliposomes. Moreover, the extracellular domain of 4F2hc suffices to stabilize solubilized LAT2. The interaction of 4F2hc with LAT2 gives insights into the structural bases for light subunit recognition and the stabilizing role of the ancillary protein in HATs.

Keywords: 4F2hc ectodomain; CD98hc.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adaptor Proteins, Signal Transducing / chemistry*
  • Adaptor Proteins, Signal Transducing / metabolism*
  • Blotting, Western
  • Chromatography, Affinity
  • Chromatography, Gel
  • Fusion Regulatory Protein 1, Heavy Chain / chemistry*
  • Fusion Regulatory Protein 1, Heavy Chain / metabolism*
  • Humans
  • Microscopy, Electron, Transmission
  • Models, Molecular*
  • Pichia
  • Protein Binding
  • Protein Conformation*

Substances

  • Adaptor Proteins, Signal Transducing
  • Fusion Regulatory Protein 1, Heavy Chain
  • LAT2 protein, human
  • SLC3A2 protein, human