title: The folding of an immunoglobulin-like Greek key protein is defined by a common-core nucleus and regions constrained by topology.
abstract: TNfn3, the third fibronectin type III domain of human tenascin, is an immunoglobulin-like protein that is a good model for experimental and theoretical analyses of Greek key folding. The third fibronectin type III domain of human tenascin folds and unfolds in a two-state fashion over a range of temperature and pH values, and in the presence of stabilising salts. Here, we present a high resolution protein engineering analysis of the single rate determining transition state. The 48 mutations report on the contribution of side-chains at 32 sites in the core and loop regions. Three areas in the protein exhibit high Phi-values, indicating that they are partially structured in the transition state. First, a common-core ring of four positions in the central strands B, C, E and F, that are in close contact, form a nucleus of tertiary interactions. The two other regions that appear well-formed are the C' region and the E-F loop. The Phi-values gradually decrease away from these regions such that the very ends of the two terminal strands A and G, have Phi-values of zero. We propose a model for the folding of immunoglobulin-like proteins in which the common-core "ring" forms the nucleus for folding, whilst the C' and E-F regions are constrained by topology to pack early. Folding characteristics of a group of structurally related proteins appear to support this model.
authors.affiliation: MRC Centre Protein Engineering, University Chemical Laboratory, Lensfield Road, Cambridge, CB2 1EW, UK.
authors.forename: S J
authors.initials: SJ
authors.lastname: Hamill
authors.forename: A
authors.initials: A
authors.lastname: Steward
authors.forename: J
authors.initials: J
authors.lastname: Clarke
mesh_terms: D020816:Amino Acid Motifs; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007136:Immunoglobulins / Q000737:chemistry*; D007700:Kinetics; D008958:Models, Molecular; D016297:Mutagenesis, Site-Directed / Q000235:genetics; D009154:Mutation / Q000235:genetics; D010446:Peptide Fragments / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D011489:Protein Denaturation; D017510:Protein Folding; D020673:Protein Renaturation; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D019063:Tenascin / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D013816:Thermodynamics
chemical_list: D007136:Immunoglobulins; D010446:Peptide Fragments; D019063:Tenascin; C114731:tenascin fibronectin type III domain (1-92)