Folded-back solution structure of monomeric factor H of human complement by synchrotron X-ray and neutron scattering, analytical ultracentrifugation and constrained molecular modelling

Factor H (FH) is a regulatory cofactor for the protease factor I in the bre akdown of C3b in the complement system of immune defence, and binds to hepa rin and other polyanionic substrates. FH is composed of 20 short consensus/ complement repeat (SCR) domains, for which the overall arrangement in solut ion is unknown. As previous studies had shown that FH can form monomeric or dimeric structures, X-ray and neutron scattering was accordingly performed with FH in the concentration range bet-M een 0.7 and 14 mg ml(-1) The radi us of gyration of FH was determined to be 11.1-11.3 nm by both methods, and the radii of gyration of the cross-section were 4.4 nm and 1.7 nm. The dis tance distribution function P(r) showed that the overall length of FH was 3 8 nm. The neutron data showed that FH was monomeric with a molecular mass o f 165,000(+/- 17,000) Da. Analytical ultracentrifugation data confirmed thi s, where sedimentation equilibrium curve fits gave a mean molecular mass of 155,000(+/-3,000) Da. Sedimentation velocity experiments using the g*(s) d erivative method showed that FH was monodisperse and had a sedimentation co efficient of 5.3(+/-0.1) S. In order to construct a full model of FH for sc attering curve and sedimentation coefficient fits, homology models were con structed for 17 of the 20 SCR domains using knowledge of the NMR structures for FH SCR-5, SCR-15 and SCR-16, and vaccinia coat protein SCR-3 and SCR-4 . Molecular dynamics simulations were used to generate a large conformation al library for each of the 19 SCR-SCR linker peptides. Peptides from these libraries were combined with the 20 SCR structures in order to generate ste reochemically complete models for the FH structure. Using an automated cons trained fit procedure, the analysis of 16,752 possible FH models showed tha t only those models in which the 20 SCR domains were bent back upon themsel ves were able to account for the scattering and sedimentation data. The bes t-fit models showed that FH had an overall length of 38 nm and is flexible. This length is significantly less than a predicted length of 73 nm if the 20 SCR structures had been arranged in an extended arrangement. This outcom e is attributed to several long linker sequences. These bent-back domain st ructures may correspond to conformational flexibility in FH and enable the multiple FH binding sites for C3 and heparin to come into close proximity. (C) 2001 Academic Press.