title: C-reactive protein exists in an NaCl concentration-dependent pentamer-decamer equilibrium in physiological buffer.
abstract: C-reactive protein (CRP) is an acute phase protein of the pentraxin family that binds ligands in a Ca(2+)-dependent manner, and activates complement. Knowledge of its oligomeric state in solution and at surfaces is essential for functional studies. Analytical ultracentrifugation showed that CRP in 2 mM Ca(2+) exhibits a rapid pentamer-decamer equilibrium. The proportion of decamer decreased with an increase in NaCl concentration. The sedimentation coefficients s(20,w)(0) of pentameric and decameric CRP were 6.4 S and in excess of 7.6 S, respectively. In the absence of Ca(2+), CRP partially dissociates into its protomers and the NaCl concentration dependence of the pentamer-decamer equilibrium is much reduced. By x-ray scattering, the radius of gyration R(G) values ranged from 3.7 nm for the pentamer to above 4.0 nm for the decamer. An averaged K(D) value of 21 microM in solution (140 mM NaCl, 2 mM Ca(2+)) was determined by x-ray scattering and modeling based on crystal structures for the pentamer and decamer. Surface plasmon resonance showed that CRP self-associates on a surface with immobilized CRP with a similar K(D) value of 23 microM (140 mM NaCl, 2 mM Ca(2+)), whereas CRP aggregates in low salt. It is concluded that CRP is reproducibly observed in a pentamer-decamer equilibrium in physiologically relevant concentrations both in solution and on surfaces. Both 2 mM Ca(2+) and 140 mM NaCl are essential for the integrity of CRP in functional studies and understanding the role of CRP in the acute phase response.
authors.affiliation: Department of Structural and Molecular Biology, Darwin Building, University College London, Gower Street, London WC1E 6BT, United Kingdom.
authors.forename: Azubuike I
authors.initials: AI
authors.lastname: Okemefuna
authors.forename: Lasse
authors.initials: L
authors.lastname: Stach
authors.forename: Sudeep
authors.initials: S
authors.lastname: Rana
authors.forename: Akim J Ziai
authors.initials: AJ
authors.lastname: Buetas
authors.forename: Jayesh
authors.initials: J
authors.lastname: Gor
authors.forename: Stephen J
authors.initials: SJ
authors.lastname: Perkins
mesh_terms: D000210:Acute-Phase Reaction / Q000378:metabolism; D002021:Buffers; D002097:C-Reactive Protein / Q000737:chemistry* / Q000378:metabolism; D002118:Calcium / Q000737:chemistry* / Q000378:metabolism; D006801:Humans; D055503:Protein Multimerization / Q000502:physiology*; D020836:Protein Structure, Quaternary / Q000502:physiology; D012965:Sodium Chloride / Q000737:chemistry* / Q000378:metabolism
chemical_list: D002021:Buffers; D012965:Sodium Chloride; D002097:C-Reactive Protein; D002118:Calcium