title: Complement factor H binds to denatured rather than to native pentameric C-reactive protein.
abstract: Binding of the complement regulatory protein, factor H, to C-reactive protein has been reported and implicated as the biological basis for association of the H402 polymorphic variant of factor H with macular degeneration. Published studies utilize solid-phase or fluid-phase binding assays to show that the factor H Y402 variant binds C-reactive protein more strongly than H402. Diminished binding of H402 variant to C-reactive protein in retinal drusen is posited to permit increased complement activation, driving inflammation and pathology. We used well validated native human C-reactive protein and pure factor H Y402H variants to test interactions. When factor H variants were incubated with C-reactive protein in the fluid phase at physiological concentrations, no association occurred. When C-reactive protein was immobilized on plastic, either non-specifically by adsorption in the presence of Ca(2+) to maintain its native fold and pentameric subunit assembly or by specific Ca(2+)-dependent binding to immobilized natural ligands, no specific binding of either factor H variant from the fluid phase was observed. In contrast, both factor H variants reproducibly bound to C-reactive protein immobilized in the absence of Ca(2+), conditions that destabilize the native fold and pentameric assembly. Both factor H variants strongly bound C-reactive protein that was denatured by heat treatment before immobilization, confirming interaction with denatured but not native C-reactive protein. We conclude that the reported binding of factor H to C-reactive protein results from denaturation of the C-reactive protein during immobilization. Differential binding to C-reactive protein, thus, does not explain association of the Y402H polymorphism with macular degeneration.
authors.affiliation: Department of Medical Biochemistry and Immunology and Pharmacology, School of Medicine, Cardiff University, Cardiff CF14 4XN, UK.
authors.forename: Svetlana
authors.initials: S
authors.lastname: Hakobyan
authors.forename: Claire L
authors.initials: CL
authors.lastname: Harris
authors.forename: Carmen W
authors.initials: CW
authors.lastname: van den Berg
authors.forename: Maria Carmen
authors.initials: MC
authors.lastname: Fernandez-Alonso
authors.forename: Elena Goicoechea
authors.initials: EG
authors.lastname: de Jorge
authors.forename: Santiago Rodriguez
authors.initials: SR
authors.lastname: de Cordoba
authors.forename: German
authors.initials: G
authors.lastname: Rivas
authors.forename: Palma
authors.initials: P
authors.lastname: Mangione
authors.forename: Mark B
authors.initials: MB
authors.lastname: Pepys
authors.forename: B Paul
authors.initials: BP
authors.lastname: Morgan
mesh_terms: D019943:Amino Acid Substitution; D001665:Binding Sites / Q000235:genetics; D002097:C-Reactive Protein / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D002118:Calcium / Q000737:chemistry / Q000378:metabolism; D017242:Complement Factor H / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D006801:Humans; D008268:Macular Degeneration; D011110:Polymorphism, Genetic; D011485:Protein Binding / Q000235:genetics; D011489:Protein Denaturation / Q000235:genetics; D017510:Protein Folding; D020836:Protein Structure, Quaternary / Q000235:genetics
chemical_list: D017242:Complement Factor H; D002097:C-Reactive Protein; D002118:Calcium
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