Complement Inhibitor of C5 Activation from the Soft Tick Ornithodoros moubata

Abstract

Blood-feeding ticks must control C activation or be damaged by the host inflammatory response. We report the characterization and expression of a novel, relatively small, broad-acting C inhibitory protein (termed OmCI) from the soft tick Ornithodoros moubata. The native 17-kDa nonglycosylated protein inhibits both human and guinea pig classical and alternative C activation pathways. The IC50 values for each pathway were 12 and 27 nM, respectively, in hemolytic assays using human serum diluted 40-fold. The cDNA encodes a protein of 168 aa, including an 18-aa secretion signal sequence that is absent in the mature form. The inhibitor has 46% amino acid identity with moubatin, a platelet aggregation inhibitor also from O. moubata that is an outlying member of the lipocalin family. Native OmCI had no inhibitory effect on the addition of C8 and C9 to preformed C5b-C7 and C5b-C8 to form the membrane attack complex and no effect on the rate of C3a production by the C3 convertase enzymes C4bC2a, C3(H2O)Bb, or C3bBb. Both recombinant and native OmCI abolish production of C5a by human classical (C4bC3bC2a) and alternative (C3bC3bBb) C5 convertases. Addition of excess C5 but not C3 competes away the inhibitory activity of OmCI, indicating that OmCI targets C5 itself rather than inhibiting the C5 convertase C4bC3bC2a itself. Direct binding of OmCI to C5 was demonstrated by Western blotting and gel filtration chromatography using 125I-labeled proteins. OmCI is the first lipocalin family member shown to inhibit C and also the first natural inhibitor that specifically targets the C5 activation step.

Combined Inhibition of Complement (C5) and CD14 Markedly Attenuates Inflammation, Thrombogenicity, and Hemodynamic Changes in Porcine Sepsis

“…This highlights the close relationship between inflammation and coagulation and shows a potent and remarkable effect of upstream inhibition of PRRs. OmCI binds directly to C5 and prevents cleavage by the C5 convertases, thus preventing formation of C5a and TCC (28,38). The attenuating effect on TF is in accordance with previous reports describing C5a-dependent TF expression in both neutrophils and endothelial cells (15,16).…”

An insight into the sialome of the soft tick, Ornithodorus parkeri

“…Remarkably, while lipocalins in general are binders of hydrophobic ligands, in all the cases mentioned above they bind hydrophilic molecules. In addition to these small ligand binding activity, some lipocalins also inhibit blood coagulation Ribeiro et al, 1995;Zhang et al, 1998), platelet aggregation (Keller et al, 1993;Waxman and Connolly, 1993), alternative complement activation (Nunn et al, 2005), and have toxic properties (Mans et al, 2003;Mans et al, 2002c). Despite this accumulating functional knowledge on hematophagous arthropods' salivary lipocalins, the vast majority of salivary lipocalins have unknown function.…”

“…OP-24, OP-6 and OP-540, belonging to clade 4 are also abundantly transcribed (17, 7 and 6 and ESTs, respectively), and were all identified in the proteomic experiments. Although lipocalins and other protein families, such as the odorant binding protein family, are known or "expected" to bind small ligands in the barrels they form, they are also known, as described above, to have additional functions such as anti-clotting Isawa et al, 2002) and anticomplement functions (Nunn et al, 2005) that do not reside in their conventional binding pocket, but rather on the protein's outer surface. Accordingly, it is possible that the remaining lipocalins not only may bind small compounds that are physiologically active at lower concentrations, such as prostaglandins and leukotrienes (Calvo et al, 2006), but they may also bind larger cytokines and chemokines.…”

Complement evasion by human pathogens

“…Finally, soft ticks of the genus Ornithodoros express at least one protein that keeps complement activation in check during their blood meal. The 17 kDa Ornithodoros moubata complement inhibitor has been shown to directly bind to C5 and prevent its cleavage, probably by inhibiting substrate binding to the C5 convertase 50,51 .…”