Relevance of lipoproteins, membranes, and extracellular vesicles in understanding C-reactive protein biochemical structure and biological activities

Abstract

Early purification protocols for C-reactive protein (CRP) often involved co-isolation of lipoproteins, primarily very low-density lipoproteins (VLDLs). The interaction with lipid particles was initially attributed to CRP’s calcium-dependent binding affinity for its primary ligand—phosphocholine—the predominant hydrophilic head group expressed on phospholipids of most lipoprotein particles. Later, CRP was shown to additionally express binding affinity for apolipoprotein B (apo B), a predominant apolipoprotein of both VLDL and LDL particles. Apo B interaction with CRP was shown to be mediated by a cationic peptide sequence in apo B. Optimal apo B binding required CRP to be surface immobilized or aggregated, treatments now known to structurally change CRP from its serum soluble pentamer isoform (i.e., pCRP) into its poorly soluble, modified, monomeric isoform (i.e., mCRP). Other cationic ligands have been described for CRP which affect complement activation, histone bioactivities, and interactions with membranes. mCRP, but not pCRP, binds cholesterol and activates signaling pathways that activate pro-inflammatory bioactivities long associated with CRP as a biomarker. Hence, a key step to express CRP’s biofunctions is its conversion into its mCRP isoform. Conversion occurs when (1) pCRP binds to a membrane surface expressed ligand (often phosphocholine); (2) biochemical forces associated with binding cause relaxation/partial dissociation of secondary and tertiary structures into a swollen membrane bound intermediate (described as mCRPm or pCRP*); (3) further structural relaxation which leads to total, irreversible dissociation of the pentamer into mCRP and expression of a cholesterol/multi-ligand binding sequence that extends into the subunit core; (4) reduction of the CRP subunit intrachain disulfide bond which enhances CRP’s binding accessibility for various ligands and activates acute phase proinflammatory responses. Taken together, the biofunctions of CRP involve both lipid and protein interactions and a conformational rearrangement of higher order structure that affects its role as a mediator of inflammatory responses.

Fueling the flames of colon cancer – does CRP play a direct pro-inflammatory role?

“…In our cohort, we found that mCRP, in addition to the above-described distribution pattern, often was scattered diffusely as small granules embedded within the stroma, unrelated to any particular cell type. Consistent with previous studies delineating the precise ligands for mCRP (5), this morphological pattern could indicate possible crosstalk between mCRP and components of the ECM. Given the putative pro-inflammatory properties of mCRP, such direct interactions could potentially contribute to excessive stromal formation.…”

“…Recently, evidence has been advanced showing that CRP exists in different structural isoforms with distinct biological activities (4). The circulating CRP isoform is a highly soluble pentameric molecule (pCRP) composed of 5 identical globular subunits arranged in a ringshaped structure (5). Each subunit contains a calcium dependent binding site enabling interaction with phosphocholine (PC), a major component of plasma membranes, defined as the primary ligand for pCRP.…”

A biofunctional review of C-reactive protein (CRP) as a mediator of inflammatory and immune responses: differentiating pentameric and modified CRP isoform effects

“…Adhesion was negated in the presence of an anti-CD18 mAb, correlating with the previous finding that pCRP with anti-P-selectin and anti-CD18 promoted complete blockade of adhesion between neutrophils and monocytes (See Neutrophil Discussion) (65). mCRP (1-30 mg/mL), specifically, also increased monocyte chemoattractant protein-1 (MCP-1) and IL-8 secretion, key mediators of leukocyte recruitment, as well as neutrophilendothelial cell adhesion via the expression of ICAM-1, VCAM-1 and E-selectin (25). To observe the similar effects with nCRP preparations, a 6x longer incubation was required, implicating that pentameric dissociation had occurred.…”

“…Hydrophobic interactions with lipid rafts are predicted to compensate for non-receptor binding. Evidence for lipid-promoted conformational dynamics is exhibited by the association of mCRP with extracellular lipid vesicles ( 26 ). Activated leukocytes slough membrane-bound CRP, which enters circulation as extracellular vesicles upon cleavage.…”

“…Circulating pCRP localizes to sites of damaged tissue (infection/injury) prior to undergoing a conformational change upon binding to damaged membranes, which exposes the proinflammatory CRP neoepitope of amino acids 199-206 (21, 22). Binding of pCRP to damaged membranes is posited to occur via lysophosphatidylcholine (LPC), a bioactive lipid found on the immune cell surface (23)(24)(25). Direct binding between the neoepitope of pCRP*/mCRP and membrane receptors is suggested to occur via FcgRIIIa (CD16), but this binding event only partially explains mCRP's adhesive properties.…”