Transient 2D IR spectroscopy of ubiquitin unfolding dynamics.
Transient two-dimensional infrared (2D IR) spectroscopy is used as a probe of protein unfolding dynamics in a direct comparison of fast unfolding experiments with molecular dynamics simulations. In the experiments, the unfolding of ubiquitin is initiated by a laser temperature jump, and protein structural evolution from nanoseconds to milliseconds is probed using amide I 2D IR spectroscopy. The temperature jump prepares a subensemble near the unfolding transition state, leading to quasi-barrierless unfolding (the "burst phase") before the millisecond activated unfolding kinetics. The burst phase unfolding of ubiquitin is characterized by a loss of the coupling between vibrations of the beta-sheet, a process that manifests itself in the 2D IR spectrum as a frequency blue-shift and intensity decrease of the diagonal and cross-peaks of the sheet's two IR active modes. As the sheet unfolds, increased fluctuations and solvent exposure of the beta-sheet amide groups are also characterized by increases in homogeneous linewidth. Experimental spectra are compared with 2D IR spectra calculated from the time-evolving structures in a molecular dynamics simulation of ubiquitin unfolding. Unfolding is described as a sequential unfolding of strands in ubiquitin's beta-sheet, using two collective coordinates of the sheet: (i) the native interstrand contacts between adjacent beta-strands I and II and (ii) the remaining beta-strand contacts within the sheet. The methods used illustrate the general principles by which 2D IR spectroscopy can be used for detailed dynamical comparisons of experiment and simulation.
Chung
Hoi Sung
HS
Department of Chemistry, Massachusetts Institute of Technology, Cambridge, MA 02139, USA.
Ganim
Ziad
Z
Jones
Kevin C
KC
Tokmakoff
Andrei
A
D003198:Computer Simulation; D018360:Crystallography, X-Ray; D008958:Models, Molecular; D011489:Protein Denaturation; D017510:Protein Folding; D017434:Protein Structure, Tertiary; D013055:Spectrophotometry, Infrared / Q000379:methods*; D013696:Temperature; D025801:Ubiquitin / Q000737:chemistry* / Q000378:metabolism*
D025801:Ubiquitin