Figure 4
From: A haplotype in CFH family genes confers high risk of rare glomerular nephropathies
The CFHR3 T risk allele of rs61737525 results decreased binding affinity to C3b and a weaker fluid-phase cofactor activity. C3b was immobilized to a CM5 chip using standard amine coupling. Duplicate injections of CFHR3WT and CFHR3Arg142Cys were performed (concentrations of 10–300 ug/ml) in 10 mM Hepes-buffered saline with 3 mM ENTA and 0.05% (vol/vol) surfactant p20. Overlaid sensograms of binding of (A) CFHR3WT and (B) CFHR3Arg142Cys to C3b, showing a kinetic and steady-state response, respectively. (C) The CFHR3Arg142Cys exhibits a binding to C3b (KD = 1.8 × 10−5M; Rmax = 194.4RU; chi-squared = 0.47), which was lower than that of CFHR3WT (KD = 1.5 × 10−7M; Rmax = 37.2RU; chi-squared = 8.81). (D) Limiting concentrations (700 ug/ml) of CFHR3WT and CFHR3Arg142Cys were incubated with factor I and the substrate C3b with increasing time points. Subsequently, the loss of the intact C3b α’-chain and appearance of its factor I cleavage products (43-kDa and 68-kDa fragments) were run on Western blot and visualized. It is apparent that mutant (Arg142Cys) CFHR3 blocks production of cleavage fragments compared to the wild-type one.
