Fig. 6: MFHR13 binds to C3b and exhibits cofactor and decay acceleration activities in fluid phase.
From: A synthetic protein as efficient multitarget regulator against complement over-activation
a C3b binding was evaluated by ELISA. Nonlinear regression using the sigmoidal equation (dose–response curve, 4 parameters) and comparison of IC50 showed no significant differences between MFHR13 and MFHRI62 binding to C3b (P = 0.8426). Data represent mean values ± SD from 5 independent experiments. Absorbance values from one representative experiment are shown in Supplementary Fig. 8b. b Cofactor activity of FI-mediated proteolytic cleavage of C3b α′-chain in the presence of MFHR1I62, FH, or MFHR13, evaluated in increasing reaction times. Densitometric analysis of C3b cleavage. The amount of intact α′-chain was normalized for each sample with the β-chain and set to 100% intact C3b α′-chain at time zero. The data represent mean value from 5 or 6 independent experiments ± SD. c One representative experiment included in b is shown. C3b, FI, and MFHR1I62, MFHR13 or FH were incubated for 10 and 20 min and C3b cleavage was visualized by SDS-PAGE and Coomassie staining. d MFHR13 displaced C3 convertases more efficiently than MFHR1I62 and MFHR1V62. C3bBb were assembled in C3b-coated microtiter plates with FB and FD. Complement regulators were added and the dissociation of the C3 convertases was detected by the amount of FB using polyclonal anti-FB antibodies. Absorbance of samples without regulator was set to 100% FB and samples without FD as the negative control. Nonlinear regression using the sigmoidal equation and comparison of IC50 indicated significant differences in DAA between MFHR13, MFHR1V62, and MFHR1I62 (P < 0.0001 for each comparison) and not significant difference between MFHR13 and hFH (P = 0.2022). The data represent mean value from 2 independent measurements ± SD.
