title: Cooperative interactions and a non-native buried Trp in the unfolded state of an SH3 domain.
abstract: The presence of residual structure in the unfolded state of the N-terminal SH3 domain of Drosophila drk (drkN SH3 domain) has been investigated using far- and near-UV circular dichroism (CD), fluorescence, and NMR spectroscopy. The unfolded (U(exch)) state of the drkN SH3 domain is significantly populated and exists in equilibrium with the folded (F(exch)) state under non-denaturing conditions near physiological pH. Denaturation experiments have been performed on the drkN SH3 domain in order to monitor the change in ellipticity, fluorescence intensity, and chemical shift between the U(exch) state and chemically or thermally denatured states. Differences between the unfolded and chemically or thermally denatured states highlight specific areas of residual structure in the unfolded state that are cooperatively disrupted upon denaturation. Results provide evidence for cooperative interactions in the unfolded state involving residues of the central beta-sheet, particularly the beta4 strand. Denaturation as well as hydrogen-exchange experiments demonstrate a non-native burial of the Trp ring within this "cooperative" core of the unfolded state. These findings support the presence of non-native hydrophobic clusters, organised by Trp rings, within disordered states.
authors.affiliation: Department of Structural Biology and Biochemistry, The Hospital for Sick Children, Toronto, Ont., Canada.
authors.forename: Karin A
authors.initials: KA
authors.lastname: Crowhurst
authors.forename: Martin
authors.initials: M
authors.lastname: Tollinger
authors.forename: Julie D
authors.initials: JD
authors.lastname: Forman-Kay
mesh_terms: D000494:Allosteric Regulation; D000495:Allosteric Site; D000595:Amino Acid Sequence; D002942:Circular Dichroism; D029721:Drosophila Proteins / Q000737:chemistry* / Q000378:metabolism*; D019791:Guanidine / Q000494:pharmacology; D006859:Hydrogen / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D057927:Hydrophobic and Hydrophilic Interactions; D009682:Magnetic Resonance Spectroscopy; D008958:Models, Molecular; D008969:Molecular Sequence Data; D011489:Protein Denaturation / Q000187:drug effects; D017510:Protein Folding; D013050:Spectrometry, Fluorescence; D013696:Temperature; D013816:Thermodynamics; D014364:Tryptophan / Q000378:metabolism*; D018909:src Homology Domains / Q000187:drug effects
chemical_list: D029721:Drosophila Proteins; C080530:drk protein, Drosophila; D006859:Hydrogen; D014364:Tryptophan; D019791:Guanidine