Figure 5
Interaction of R139W withfactorB. (A) R139 position on the structures of C3b with FB in closed (refractory to cleavage by FD) and open (prone to cleavage by FD) conformations and on the structure of the C3bBb complex. The α and β chains of C3b are depicted in blue and green and FB is colored in magenta. (B) Binding of FB to WT or mutant recombinant C3, bound to an anti-C3d mAb, coated to the ELISA plate. Serum-free supernatant was used as a source of recombinant C3 molecules. (C) The binding of FB to recombinant WT or R139W C3, bound to an anti-C3d mAb on the CM5 biosensor chip, was studied by SPR using Biacore. Recombinant C3 proteins, purified from serum-free culture supernatants by DEAE-Sepharose column, were used. (D) Formation of a C3 convertase on the biosensor chip by subsequent injection of C3+FB+FD, followed by an injection of FB+FD. Purified WT or R139W recombinant C3 were mixed with native C3, purified from plasma. Proteins deposition on the chip was followed over time. For panels B through D, 1 representative experiment of 3, performed with 3 independent productions of C3, is presented.

Interaction of R139W withfactorB. (A) R139 position on the structures of C3b with FB in closed (refractory to cleavage by FD) and open (prone to cleavage by FD) conformations and on the structure of the C3bBb complex. The α and β chains of C3b are depicted in blue and green and FB is colored in magenta. (B) Binding of FB to WT or mutant recombinant C3, bound to an anti-C3d mAb, coated to the ELISA plate. Serum-free supernatant was used as a source of recombinant C3 molecules. (C) The binding of FB to recombinant WT or R139W C3, bound to an anti-C3d mAb on the CM5 biosensor chip, was studied by SPR using Biacore. Recombinant C3 proteins, purified from serum-free culture supernatants by DEAE-Sepharose column, were used. (D) Formation of a C3 convertase on the biosensor chip by subsequent injection of C3+FB+FD, followed by an injection of FB+FD. Purified WT or R139W recombinant C3 were mixed with native C3, purified from plasma. Proteins deposition on the chip was followed over time. For panels B through D, 1 representative experiment of 3, performed with 3 independent productions of C3, is presented.

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