Structural basis for the reduced affinity of mfH with fHbp.

<p>(<b>A</b>) Cartoon of hfH₆₇ viewed from through V1 fHbp (solid line) with amino acids changed in hfH with murine residues (outlined by yellow dashes), and those replaced in mfH with human residues (outlined by light blue dashes). (<b>B</b>) SPR analysis of binding of two hfH₆₇ mutants each containing two amino acid changes (shown) with fHbps from each variant family. (<b>C</b>) Far western analysis of V1 fHbp and a control protein, PPX; blots were overlaid with 5 µg/ml of the recombinant proteins mfH, modified mfH (with 14 humanised amino acids) or hfH, or with human serum (1 in 2000 dilution) as indicated; the sizes of the mol. wt. marker are shown. (<b>D</b>) Structure of mfH₆₇ (blue ribbon) superimposed on V1 fHbp (white ribbon) and hfH (green ribbon). While fH₆ from both species are superimposable, the orientation of fH₇ differs significantly between mfH and hfH (indicated in red dashed circle).</p

Non-functional fHbps retain their immunogenicity in transgenic mice.

<p>(<b>A</b>) ELISAs assaying anti-V1 titres elicited in pooled sera following immunisation of transgenic mice with the wild-type protein and non-functional V1 fHbps. (<b>B</b>) SBA titres of sera from individual mice immunisation with fHbps.</p

Effect of mutations at positions equivalent to fHbp V1 residues 283 and 304 (<i>i.e.</i> Thr³⁰⁴ in V2 and V3 fHbp) on the <i>K</i>_D for binding to fH₆₇, shown relative to the wild-type proteins.

<p>DM indicates double Ala substitution; ND, not determined.</p