Carbon-Detected (15)N NMR Spin Relaxation of an Intrinsically Disordered Protein: FCP1 Dynamics Unbound and in Complex with RAP74.
Intrinsically disordered proteins (IDPs) lack unique 3D structures under native conditions and as such exist as highly dynamic ensembles in solution. We present two (13)C-direct detection experiments for the measurement of (15)N NMR spin relaxation called the CON(T1)-IPAP and CON(T2)-IPAP that quantify backbone dynamics on a per-residue basis for IDPs in solution. These experiments have been applied to the intrinsically disordered C-terminal of FCP1, both free in solution and while bound to the RAP74 winged-helix domain. The results provide evidence that most of FCP1 remains highly dynamic in both states, while the 20 residues forming direct contact with RAP74 become more ordered in the complex. Parallel analysis of RAP74 backbone (15)N NMR spin relaxation reveals only very limited ordering of RAP74 upon FCP1 binding. Taken together, these data show that folding-upon-binding is highly local in this system, with disorder prevailing even in the complex.
Department of Chemistry, The Pennsylvania State University, 104 Chemistry Building, University Park, Pennsylvania 16802, United States.
Chad W
CW
Lawrence
Department of Chemistry, The Pennsylvania State University, 104 Chemistry Building, University Park, Pennsylvania 16802, United States.
Scott A
SA
Showalter
NMR; intrinsically disordered protein; protein−protein interaction; spin relaxation; transactivation domain