Mapping of the binding of platelet-derived growth factor to distinct domains of the basement membrane proteins BM-40 and perlecan and distinction from the BM-40 collagen-binding epitope

Eur J Biochem. 1998 Jul 1;255(1):60-6. doi: 10.1046/j.1432-1327.1998.2550060.x.

Abstract

A surface plasmon resonance assay was used to analyze the binding of platelet-derived growth factor (PDGF)-AA and PDGF-BB to various proteins of the extracellular matrix. This identified several collagen types; laminin-1, nidogen, perlecan and BM-40 as potential ligands for PDGF with Kd values in the range 2-3200 nM. Perlecan and BM-40 were used to examine the domain specificity and other parameters of the interactions. Recombinant human and mouse BM-40 were shown to bind both PDGFs in a similar manner with a Kd of about 5-10 nM. Studies with deletion mutants of human BM-40 demonstrated binding to its C-terminal extracellular calcium-binding (EC) module, yet the interaction did not require calcium. This distinguishes this from the binding of the EC module to various collagen types, which is strictly calcium dependent. Furthermore, deletion of helix alphaC or two point mutations in helix alphaA of the EC module either enhanced or abolished binding to collagen IV. Since these mutations had no effects on binding to PDGF, it demonstrated the presence of two different binding epitopes. Binding of PDGF-BB to the perlecan core protein could be mapped to its domain III-2 (Kd = 8 nM) with lower affinities shown for domains I, IV-1 and V (Kd = 34-64 nM). Other perlecan domains (II, III-1, III-3, IV-2) were inactive. PDGF-AA was also shown to bind domain III-2 but not III-1. Neither nidogen, BM-40 or perlecan domain III-2 interfered with the binding of PDGF to its alpha and beta receptors, however, suggesting that these interactions may be mainly used for storage of PDGF in the extracellular matrix.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Basement Membrane
  • Becaplermin
  • Binding Sites
  • Biosensing Techniques
  • Collagen / metabolism
  • DNA Mutational Analysis
  • Extracellular Matrix Proteins / metabolism*
  • Heparan Sulfate Proteoglycans*
  • Heparitin Sulfate / metabolism*
  • Humans
  • Mice
  • Osteonectin / genetics
  • Osteonectin / metabolism*
  • Platelet-Derived Growth Factor / metabolism*
  • Protein Binding
  • Proteoglycans / metabolism*
  • Proto-Oncogene Proteins c-sis
  • Receptors, Platelet-Derived Growth Factor / metabolism
  • Recombinant Proteins / metabolism
  • Swine

Substances

  • Extracellular Matrix Proteins
  • Heparan Sulfate Proteoglycans
  • Osteonectin
  • Platelet-Derived Growth Factor
  • Proteoglycans
  • Proto-Oncogene Proteins c-sis
  • Recombinant Proteins
  • platelet-derived growth factor A
  • perlecan
  • Becaplermin
  • Collagen
  • Heparitin Sulfate
  • Receptors, Platelet-Derived Growth Factor