Complement Factor H Binds to Denatured Rather than to Native Pentameric C-reactive Protein

• Published in: The Journal of biological chemistry 2008-11, Vol.283 (45), p.30451-30460
• Main Authors: Hakobyan, Svetlana, Harris, Claire L., van den Berg, Carmen W., Fernandez-Alonso, Maria Carmen, de Jorge, Elena Goicoechea, de Cordoba, Santiago Rodriguez, Rivas, German, Mangione, Palma, Pepys, Mark B., Morgan, B. Paul
• Format: Article
• Language: English
• Subjects: Amino Acid Substitution; Binding Sites - genetics; C-Reactive Protein - chemistry; C-Reactive Protein - genetics; C-Reactive Protein - metabolism; Calcium - chemistry; Calcium - metabolism; Complement Factor H - chemistry; Complement Factor H - genetics; Complement Factor H - metabolism; Humans; Macular Degeneration; Polymorphism, Genetic; Protein Binding - genetics; Protein Denaturation - genetics; Protein Folding; Protein Structure and Folding; Protein Structure, Quaternary - genetics
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1074/jbc.M803648200

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 Ca2+ to maintain its native fold and pentameric subunit assembly or by specific Ca2+-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 Ca2+, 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.