FIGURE 2.

Fluid-phase interaction between native CRP and fH by analytical ultracentrifugation. A, representative sedimentation equilibrium gradients (9000 rpm) for the individual variants, fH-Y402 and fH-H402 (superimposed in this dataset; closed circles), and CRP (open circles) and the mixtures of fH-Y402 and CRP (open triangles) and fH-H402 and CRP (closed triangles), all at 0.2 g/liter. The dashed line shows the calculated best fit for a hypothetical 1:1 complex for fH and CRP. Residuals for each dataset are shown in the box below. B and C, gel-filtration chromatograms from the final purification step of plasma fH Y402 and H402, respectively. The inset in B shows Coomassie staining of the final fH preparations run in PAGE. D, representative sedimentation equilibrium gradients (9000 rpm) for the individual variants fH-Y402 and fH-H402 (superimposed in this dataset; closed circles) and C3b (open circles) and the mixtures of fH-Y402 and C3b (open triangles) and fH-H402 and C3b (closed triangles), all at 0.2 g/liter. Residuals for each dataset are shown in the box below. The inset shows a sample dataset of one equilibrium sedimentation gradient for fH (open circles), C3b (open triangles), and the mixture of fH and C3b (closed circles). Theoretical lines for no interaction (dotted) and a 1:1 interaction (dashed) are shown. The solid line shows the calculated best fit for the mixture. E, gel-filtration chromatogram from the final purification step of trypsin-digested C3b; the inset shows Coomassie staining of the final C3b preparation. OD, optical density.