title: Estimation of interdomain flexibility of N-terminus of factor H using residual dipolar couplings.
abstract: Characterization of segmental flexibility is needed to understand the biological mechanisms of the very large category of functionally diverse proteins, exemplified by the regulators of complement activation, that consist of numerous compact modules or domains linked by short, potentially flexible, sequences of amino acid residues. The use of NMR-derived residual dipolar couplings (RDCs), in magnetically aligned media, to evaluate interdomain motion is established but only for two-domain proteins. We focused on the three N-terminal domains (called CCPs or SCRs) of the important complement regulator, human factor H (i.e., FH1-3). These domains cooperate to facilitate cleavage of the key complement activation-specific protein fragment, C3b, forming iC3b that no longer participates in the complement cascade. We refined a three-dimensional solution structure of recombinant FH1-3 based on nuclear Overhauser effects and RDCs. We then employed a rudimentary series of RDC data sets, collected in media containing magnetically aligned bicelles (disklike particles formed from phospholipids) under three different conditions, to estimate interdomain motions. This circumvents a requirement of previous approaches for technically difficult collection of five independent RDC data sets. More than 80% of conformers of this predominantly extended three-domain molecule exhibit flexions of <40°. Such segmental flexibility (together with the local dynamics of the hypervariable loop within domain 3) could facilitate recognition of C3b via initial anchoring and eventual reorganization of modules to the conformation captured in the previously solved crystal structure of a C3b:FH1-4 complex.
authors.affiliation: Laboratory of Molecular Biophysics, National Heart, Lung, and Blood Institute, National Institutes of Health, 50 Center Drive, Bethesda, Maryland 20892, USA.
authors.forename: Mateusz
authors.initials: M
authors.lastname: Maciejewski
authors.forename: Nico
authors.initials: N
authors.lastname: Tjandra
authors.forename: Paul N
authors.initials: PN
authors.lastname: Barlow
mesh_terms: D003167:Complement Activation; D003179:Complement C3b / Q000737:chemistry / Q000378:metabolism; D017242:Complement Factor H / Q000737:chemistry* / Q000378:metabolism; D006801:Humans; D066298:In Vitro Techniques; D008958:Models, Molecular; D056004:Molecular Dynamics Simulation; D019906:Nuclear Magnetic Resonance, Biomolecular; D054730:Protein Interaction Domains and Motifs; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism
chemical_list: D011994:Recombinant Proteins; D003179:Complement C3b; D017242:Complement Factor H