Covalent cross-linking of the Phytophthora megasperma oligopeptide elicitor to its receptor in parsley membranes

Proc Natl Acad Sci U S A. 1995 Mar 14;92(6):2338-42. doi: 10.1073/pnas.92.6.2338.

Abstract

An oligopeptide elicitor from Phytophthora megasperma f.sp. glycinea (Pep-13) that induces phytoalexin accumulation in cultured parsley cells was radioiodinated and chemically cross-linked to its binding site in microsomal and plasma membrane preparations with each of three homobifunctional reagents. Analysis by SDS/PAGE and autoradiography of solubilized membrane proteins demonstrated labeling of a 91-kDa protein, regardless of which reagent was used. Cross-linking of this protein was prevented by addition of excess unlabeled Pep-13. The competitor concentration found to half-maximally reduce the intensity of the cross-linked band was 6 nM, which is in good agreement with the IC50 value of 4.7 nM, obtained from ligand binding assays. No crosslinking of 125I-labeled Pep-13 was observed by using microsomal membranes from three other plant species, indicating species-specific occurrence of the binding site. Coupling of 125I-Pep-13 to the parsley 91-kDa protein required the same structural elements within the ligand as was recently reported for binding of 125I-Pep-13 to parsley microsomes, elicitor-induced stimulation of ion fluxes across the plasma membrane, the oxidative burst, the expression of defense-related genes, and phytoalexin production. These findings suggest that the 91-kDa protein identified in parsley membranes is the oligopeptide elicitor receptor mediating activation of a multicomponent defense response.

Publication types

  • Comparative Study
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Binding, Competitive
  • Cells, Cultured
  • Cross-Linking Reagents / pharmacology
  • Daucus carota / metabolism
  • Electrophoresis, Polyacrylamide Gel
  • Fungal Proteins / isolation & purification
  • Fungal Proteins / metabolism*
  • Glycine max / metabolism
  • Glycoproteins
  • Intracellular Membranes / metabolism
  • Iodine Radioisotopes
  • Kinetics
  • Magnoliopsida / metabolism*
  • Membrane Glycoproteins / isolation & purification
  • Membrane Glycoproteins / metabolism*
  • Membrane Proteins
  • Microsomes / metabolism
  • Molecular Sequence Data
  • Peptide Fragments / chemistry
  • Peptide Fragments / metabolism
  • Peptide Fragments / pharmacology
  • Phytoalexins
  • Phytophthora / metabolism*
  • Plant Extracts / biosynthesis
  • Sesquiterpenes
  • Substrate Specificity
  • Succinimides / pharmacology*
  • Terpenes

Substances

  • Cross-Linking Reagents
  • Fungal Proteins
  • Glycoproteins
  • Iodine Radioisotopes
  • Membrane Glycoproteins
  • Membrane Proteins
  • Peptide Fragments
  • Plant Extracts
  • Sesquiterpenes
  • Succinimides
  • Terpenes
  • elicitor protein, Phytophthora megasperma
  • dithiobis(succinimidylpropionate)
  • Phytoalexins