Sensitivity of NMR residual dipolar couplings to perturbations in folded and denatured staphylococcal nuclease.
The invariance of NMR residual dipolar couplings (RDCs) in denatured forms of staphylococcal nuclease to changes in denaturant concentration or amino acid sequence has previously been attributed to the robustness of long-range structure in the denatured state. Here we compare RDCs of the wild-type nuclease with those of a fragment that retains a folded OB-fold subdomain structure despite missing the last 47 of 149 residues. The RDCs of the intact protein and of the truncation fragment are substantially different under conditions that favor folded structure. By contrast, there is a strong correlation between the RDCs of the full-length protein and the fragment under denaturing conditions (6 M urea). The RDCs of the folded and unfolded forms of the proteins are uncorrelated. Our results suggest that RDCs are more sensitive to structural changes in folded than unfolded proteins. We propose that the greater susceptibility of RDCs in folded states is a consequence of the close packing of the polypeptide chain under native conditions. By contrast, the invariance of RDCs in denatured states is more consistent with a disruption of cooperative structure than with the retention of a unique long-range folding topology.
Sallum
Christine O
CO
Department of Molecular and Cell Biology, University of Connecticut, Storrs, Connecticut 06269-3125, USA.
Martel
David M
DM
Fournier
Robert S
RS
Matousek
William M
WM
Alexandrescu
Andrei T
AT
D016880:Anisotropy; D025561:Bacteriophage Pf1 / Q000737:chemistry; D003198:Computer Simulation; D018360:Crystallography, X-Ray; D004795:Enzyme Stability / Q000235:genetics; D029968:Escherichia coli Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D016153:Genes, Suppressor; D016014:Linear Models; D008836:Micrococcal Nuclease / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D008956:Models, Chemical; D008958:Models, Molecular; D009154:Mutation; D019906:Nuclear Magnetic Resonance, Biomolecular / Q000379:methods*; D010446:Peptide Fragments / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D011489:Protein Denaturation / Q000235:genetics; D017510:Protein Folding; D013816:Thermodynamics
D029968:Escherichia coli Proteins; D010446:Peptide Fragments; D008836:Micrococcal Nuclease