Structural basis for surface activation of the classical complement cascade by the short pentraxin C-reactive protein

Abstract

Human C-reactive protein (CRP) is a pentameric complex involved in defence against pathogens and regulation of autoimmunity. CRP is also a therapeutic target, with both administration and depletion of serum CRP being pursued as a possible treatment for autoimmune and cardiovascular diseases, among others. CRP binds to phosphocholine (PC) moieties on membranes in order to activate the complement system via the C1 complex, but it is unknown how CRP, or any pentraxin, binds to C1. Here, we present a cryo-electron tomography (cryoET)-derived structure of CRP bound to PC ligands and the C1 complex. To gain control of CRP binding, a synthetic mimotope of PC was synthesised and used to decorate cell-mimetic liposome surfaces. Structure-guided mutagenesis of CRP yielded a fully-active complex able to bind PC-coated liposomes that was ideal for cryoET and subtomogram averaging. In contrast to antibodies, which form Fc-mediated hexameric platforms to bind and activate the C1 complex, CRP formed rectangular platforms assembled from four laterally-associated CRP pentamers that bind only four of the six available globular C1 head groups. Potential residues mediating lateral association of CRP were identified from interactions between unit cells in existing crystal structures, which rationalised previously unexplained mutagenesis data regarding CRP-mediated complement activation. The structure also enabled interpretation of existing biochemical data regarding interactions mediating C1 binding, and identified additional residues for further mutagenesis studies. These structural data therefore provide a possible mechanism for regulation of complement by CRP, which limits complement progression and has consequences for how the innate immune system influences autoimmunity. Significance statementHuman C-reactive protein (CRP) activates the complement system to protect us from infections, but can also contribute towards progression of cardiovascular and autoimmune diseases when erroneously activated. To understand these processes, the authors used cryo-electron tomography to solve the in situ structure of surface-bound CRP interacting with the complement C1 complex. The structure revealed new interfaces that explain previous, sometimes contradictory, biochemical data. Comparisons with existing structures of antibody-mediated C1 activation revealed distinct structural differences that may explain how CRP modulates complement activity. Together, these structural data identify residues for mutagenesis to gain control over CRP functions, and provide new routes for future therapeutic developments.