Hemeprotein Tpx1 interacts with cell-surface heme transporter Str3 in Schizosaccharomyces pombe

Mol Microbiol. 2021 Apr;115(4):699-722. doi: 10.1111/mmi.14638. Epub 2020 Nov 21.

Abstract

Str3 is a transmembrane protein that mediates low-affinity heme uptake in Schizosaccharomyces pombe. Under iron-limiting conditions, Str3 remains at the cell surface in the presence of increasing hemin concentrations. Using a proximity-dependent biotinylation approach coupled to mass spectrometry and coimmunoprecipitation assays, we report that the peroxiredoxin Tpx1 is a binding partner of Str3. Under microaerobic conditions, cells deficient in heme biosynthesis and lacking the heme receptor Shu1 exhibit poor hemin-dependent growth in the absence of Tpx1. Analysis of membrane protein preparations from iron-starved hem1Δ shu1Δ str3Δ tpx1Δ cells coexpressing Str3-GFP and TAP-Tpx1 showed that TAP-Tpx1 is enriched in membrane protein fractions in response to hemin. Bimolecular fluorescence complementation assays brought additional evidence that an interaction between Tpx1 and Str3 occurs at the plasma membrane. Results showed that Tpx1 exhibits an equilibrium constant value of 0.26 μM for hemin. The association of Tpx1 with hemin protects hemin from degradation by H2 O2 . The peroxidase activity of hemin is lowered when it is bound to Tpx1. Taken together, these results revealed that Tpx1 is a novel interacting partner of Str3. Our data are the first example of an interaction between a cytoplasmic heme-binding protein and a cell-surface heme transporter.

Keywords: fission yeast; heme; hemoprotein; major facilitator transporter; peroxiredoxin.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Motifs
  • Biotinylation
  • Cell Membrane / metabolism
  • DNA, Fungal
  • Heme / metabolism
  • Hemeproteins / genetics
  • Hemeproteins / metabolism*
  • Hemin / metabolism
  • Hydrogen Peroxide / metabolism
  • Iron / metabolism
  • Membrane Transport Proteins / genetics*
  • Membrane Transport Proteins / metabolism*
  • Mutation
  • Oxidation-Reduction
  • Peroxiredoxins / genetics*
  • Peroxiredoxins / metabolism*
  • Protein Binding
  • Schizosaccharomyces / enzymology
  • Schizosaccharomyces / genetics*
  • Schizosaccharomyces / metabolism*
  • Schizosaccharomyces pombe Proteins / genetics*
  • Schizosaccharomyces pombe Proteins / metabolism*

Substances

  • DNA, Fungal
  • Hemeproteins
  • Membrane Transport Proteins
  • Schizosaccharomyces pombe Proteins
  • Tpx1 protein, S pombe
  • Heme
  • Hemin
  • Hydrogen Peroxide
  • Iron
  • Peroxiredoxins