Structural basis for the binding of tryptophan-based motifs by δ-COP

Proc Natl Acad Sci U S A. 2015 Nov 17;112(46):14242-7. doi: 10.1073/pnas.1506186112. Epub 2015 Nov 2.

Abstract

Coatomer consists of two subcomplexes: the membrane-targeting, ADP ribosylation factor 1 (Arf1):GTP-binding βγδζ-COP F-subcomplex, which is related to the adaptor protein (AP) clathrin adaptors, and the cargo-binding αβ'ε-COP B-subcomplex. We present the structure of the C-terminal μ-homology domain of the yeast δ-COP subunit in complex with the WxW motif from its binding partner, the endoplasmic reticulum-localized Dsl1 tether. The motif binds at a site distinct from that used by the homologous AP μ subunits to bind YxxΦ cargo motifs with its two tryptophan residues sitting in compatible pockets. We also show that the Saccharomyces cerevisiae Arf GTPase-activating protein (GAP) homolog Gcs1p uses a related WxxF motif at its extreme C terminus to bind to δ-COP at the same site in the same way. Mutations designed on the basis of the structure in conjunction with isothermal titration calorimetry confirm the mode of binding and show that mammalian δ-COP binds related tryptophan-based motifs such as that from ArfGAP1 in a similar manner. We conclude that δ-COP subunits bind Wxn(1-6)[WF] motifs within unstructured regions of proteins that influence the lifecycle of COPI-coated vesicles; this conclusion is supported by the observation that, in the context of a sensitizing domain deletion in Dsl1p, mutating the tryptophan-based motif-binding site in yeast causes defects in both growth and carboxypeptidase Y trafficking/processing.

Keywords: COPI; coatomer; membrane trafficking; vesicle coat; δ-COP μ homology domain-binding motifs.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Motifs
  • COP-Coated Vesicles / chemistry
  • COP-Coated Vesicles / genetics
  • COP-Coated Vesicles / metabolism
  • Calorimetry, Indirect
  • Cathepsin A / chemistry
  • Cathepsin A / genetics
  • Cathepsin A / metabolism
  • Coatomer Protein / chemistry*
  • Coatomer Protein / genetics
  • Coatomer Protein / metabolism
  • DNA-Binding Proteins / chemistry
  • DNA-Binding Proteins / genetics
  • DNA-Binding Proteins / metabolism
  • GTPase-Activating Proteins / chemistry
  • GTPase-Activating Proteins / genetics
  • GTPase-Activating Proteins / metabolism
  • Protein Binding
  • Protein Structure, Tertiary
  • Saccharomyces cerevisiae / chemistry*
  • Saccharomyces cerevisiae / genetics
  • Saccharomyces cerevisiae / metabolism
  • Saccharomyces cerevisiae Proteins / chemistry
  • Saccharomyces cerevisiae Proteins / genetics
  • Saccharomyces cerevisiae Proteins / metabolism
  • Tryptophan / chemistry*
  • Tryptophan / genetics
  • Tryptophan / metabolism

Substances

  • Coatomer Protein
  • DNA-Binding Proteins
  • DSL1 protein, S cerevisiae
  • GCS1 protein, S cerevisiae
  • GTPase-Activating Proteins
  • Saccharomyces cerevisiae Proteins
  • Tryptophan
  • Cathepsin A
  • PRC1 protein, S cerevisiae

Associated data

  • PDB/5FJW
  • PDB/5FJX
  • PDB/5FJZ
  • PDB/5FK0