Structural basis and functional effects of the interaction between complement inhibitor C4b-binding protein and DNA.
Human C4b-binding protein (C4BP) is a soluble, multiple-subunit inhibitor of complement that circulates in blood. Recently C4BP was shown to bind DNA, reduce DNA release from necrotic cells and limit DNA-mediated complement activation in solution. Herein we employed nuclear magnetic resonance spectroscopy to measure chemical shift perturbations and used them to restrain the computational docking of a B-form 10-base-pair DNA molecule onto the solution structure of C4BP alpha-chain complement control protein (CCP) domains 1-2 (C4BP12). Six amino acid residues located on one face of the interdomain junction - Val(38), Ser(40), Thr(43), Tyr(62), Lys(63) and Arg(64) - exhibited the largest chemical shift changes. In the model, the DNA lies in a cleft formed by the interdomain interface. The double-helix is perpendicular to the long axis of C4BP12 consistent with the multiple arms of C4BP binding to adjacent sites on a longer DNA molecule. The DNA lies in a region previously shown to bind C4b and heparin and these molecules (but not C3b) inhibited the DNA-C4BP interaction. Nonetheless, crucial C4BP functions such as cofactor activity for factor I cleavage of C4b and C3b, and decay acceleration of the classical C3 convertase appeared not to be affected by the presence of DNA. Taken together these results reinforce the case for the occupation of some of the seven arms of C4BP in a multivalent interaction with DNA or surface bound glycosaminoglycans while other arms engage C4b or C3b.
Okroj
Marcin
M
Lund University, Department of Laboratory Medicine, Section of Medical Protein Chemistry, University Hospital Malmö, Malmö, Sweden.
Jenkins
Huw T
HT
Herbert
Andrew P
AP
Barlow
Paul N
PN
Blom
Anna M
AM
D000596:Amino Acids / Q000378:metabolism; D001667:Binding, Competitive; D050577:Complement C3-C5 Convertases / Q000378:metabolism; D003179:Complement C3b / Q000378:metabolism; D015935:Complement C4b / Q000378:metabolism; D050716:Complement C4b-Binding Protein; D004247:DNA / Q000737:chemistry* / Q000378:metabolism*; D024202:Electrophoretic Mobility Shift Assay; D006493:Heparin / Q000378:metabolism; D006649:Histocompatibility Antigens / Q000737:chemistry* / Q000378:metabolism*; D006801:Humans; D008958:Models, Molecular; D011485:Protein Binding; D017433:Protein Structure, Secondary; D013329:Structure-Activity Relationship
D000596:Amino Acids; C055609:C4BPA protein, human; D050716:Complement C4b-Binding Protein; D006649:Histocompatibility Antigens; D003179:Complement C3b; D015935:Complement C4b; D006493:Heparin; D004247:DNA; D050577:Complement C3-C5 Convertases