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Functional interactions of the C-terminus of the complement regulator Factor H with its ligands

Dunne, O; (2016) Functional interactions of the C-terminus of the complement regulator Factor H with its ligands. Doctoral thesis , UCL (University College London).

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Abstract

The complement system plays a crucial role in immunity by eliminating disease-causing pathogens. Its major regulator, complement Factor H (CFH) with 20 short complement regulator (SCR) domains, prevents the destruction of host cells through excessive complement activation. CFH achieves this in the alternative pathway of complement activation through its discrimination between activator (pathogen) and non-activator (hosts) cells. Mutations and polymorphisms within CFH can lead to severe disease. Three interactions that enables CFH to protect host cells were investigated in this thesis. CFH contains a weak dimerisation site that potentially enables high concentrations of CFH to accumulate at host cell surfaces. The C-terminal SCR-16/20 domains contains an unknown dimerization site. Using seven C-terminal CFH fragments, a combination of size exclusion chromatography (SEC), analytical ultracentrifugation (AUC) and small angle X-ray scattering (SAXS) showed that this dimer site is located in SCR-17/18. Modelling suggested that the dimer was formed by an anti-parallel length-wise interaction. Because disease-causing mutations in SCR-17/18 occurred at the predicted dimer interface, this explains how these mutations may lead to disease. CFH interacts with pentameric C-reactive protein (CRP). The interaction between the seven CFH C-terminal fragments and CRP was examined to identify the CRP binding site on CFH. A combinantion of SEC, AUC, fluorescent AUC, surface plasmon resonance, microscale thermophoresis and electron microscopy revealed a weak binding site in SCR-19/20 for pentameric CRP. This interaction would allow CFH to bind to CRP at high concentrations on host cell surfaces during an acute phase response with elevated CRP plasma levels, thus protecting CRP-decorated host cells from excessive immune activation. The CFH SCR-19/20 complex with the C3d domain of C3b has been studied by crystallography to reveal either a 1:1 or 1:2 binding stoichiometry. To resolve this ambiguity, deuterium-labelled SCR-19/20 was produced to carry out contrast variation experiments by small-angle neutron scattering and SAXS. The results showed that one SCR-19/20 interacts with two C3d molecules in solution providing an extra mechanism for CFH to protect host cells during excessive inflammation.

Type: Thesis (Doctoral)
Title: Functional interactions of the C-terminus of the complement regulator Factor H with its ligands
Event: UCL (University College London)
Language: English
UCL classification: UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences
UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Life Sciences
URI: http://discovery.ucl.ac.uk/id/eprint/1493389
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