title: Biologically active decorin is a monomer in solution.
abstract: It has been reported that decorin and its protein core can have molecular masses nearly double the size of those previously published, suggesting a dimeric structure. In this study we tested whether biologically active decorin and its glycoprotein core would form dimers in solution. We used homo- and hetero-bifunctional chemical cross-linking reagents, BS3 and sulfo-SMPB, respectively, as well as glutaraldehyde and found no preferential dimer formation, whether chemical cross-linking was performed in the presence or absence of live cells. Under the same experimental conditions, we easily detected dimers of epidermal growth factor receptor and basic fibroblast growth factor, two glycoproteins known to dimerize. Only at very high cross-linker to decorin molar ratios (2000:1) were trimers and multimers observed, but performing the chemical cross-linking in the presence of a reducing agent abolished these. The elution of decorin protein core in Superose 6 gel chromatography gave masses compatible with monomeric proteins, both before and after denaturation with 2.5 M guanidine HCl. Matrix-assisted laser desorption ionization gave a mass of 44,077 Da for decorin protein core, without any evidence of dimers or oligomers. Extensive oligomerization of the decorin protein core was observed only after dialysis against water and freeze-drying. These oligomers were considered artifacts because they were independent of chemical cross-linking and were resistant to heat denaturation and disulfide-bond reduction. Oligomeric preparations showed markedly reduced biological activity in both phosphorylation and collagen fibrillogenesis assays. Thus, biologically active decorin is a monomer in solution and, as such, is a monovalent ligand for various extracellular matrix proteins, growth factors, and cell surface receptors.
authors.affiliation: Department of Pathology, Anatomy and Cell Biology, Thomas Jefferson University, Philadelphia, Pennsylvania 19107, USA.
authors.forename: Silvia
authors.initials: S
authors.lastname: Goldoni
authors.forename: Rick T
authors.initials: RT
authors.lastname: Owens
authors.forename: David J
authors.initials: DJ
authors.lastname: McQuillan
authors.forename: Zachary
authors.initials: Z
authors.lastname: Shriver
authors.forename: Ram
authors.initials: R
authors.lastname: Sasisekharan
authors.forename: David E
authors.initials: DE
authors.lastname: Birk
authors.forename: Shelly
authors.initials: S
authors.lastname: Campbell
authors.forename: Renato V
authors.initials: RV
authors.lastname: Iozzo
mesh_terms: D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D002850:Chromatography, Gel; D003432:Cross-Linking Reagents / Q000494:pharmacology; D058575:Decorin; D019281:Dimerization; D004220:Disulfides; D004305:Dose-Response Relationship, Drug; D066246:ErbB Receptors / Q000737:chemistry; D016326:Extracellular Matrix Proteins; D016222:Fibroblast Growth Factor 2 / Q000737:chemistry; D005976:Glutaral / Q000737:chemistry; D019791:Guanidine / Q000737:chemistry; D006358:Hot Temperature; D006801:Humans; D011489:Protein Denaturation; D011509:Proteoglycans / Q000737:chemistry*; D011994:Recombinant Proteins / Q000737:chemistry; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D013997:Time Factors
chemical_list: D003432:Cross-Linking Reagents; C546878:DCN protein, human; D058575:Decorin; D004220:Disulfides; D016326:Extracellular Matrix Proteins; D011509:Proteoglycans; D011994:Recombinant Proteins; D016222:Fibroblast Growth Factor 2; D066246:ErbB Receptors; D019791:Guanidine; D005976:Glutaral