Investigation of Dynamics of Villin Headpiece Subdomain by Solid-State NMR with the Use of Single-Site Labeling.
EMSL Project ID
24105
Abstract
Villin headpiece protein subdomain (HP36) is extensively used as a model system for protein folding. A mutation of F58 residue was shown to rupture the native fold. Yet a double mutant involving two other Phe residues, F47 and F51, retains the native fold but displays a decreased stability and a higher unfolding rate. (Brewer et al, Proc. Natl. Acad. Sci. U S A 2005, 102, 16662. Frank et al, Protein Sci. 2002, 11, 680.) This study proposes to address an issue of conformational heterogeneity in folded state by concentrating on the region essential for the native conformation. In the first phase of experiments, for which NMR time is requested, methyl groups of L69 will be selectively deuterated in both native HP36 and its F47F51 double mutant. The side chain of L69 is in close proximity to the essential F58 residue. Proposed solid-state NMR experiments include a combination of deuterium quadrupolar tensor line-shape analysis and relaxation techniques, such as T1 and T1r, at several magnetic field strengths and over a broad range of temperatures between 30 -285K. Comparison of the data for HP36 and the F47F51 mutant will yield detailed dynamics information in site of interest. Studies of protein dynamics on model systems such as HP36 enhance our understanding of the role of the conformational heterogeneity in the problem of protein folding. The experiments are planned to be conducted in collaboration with Dr. Paul Ellis and Dr. Andrew Lipton. Proposal Type: Biological Interactions and Dynamics, standard access, non-proprietary, general.
Project Details
Project type
Large-Scale EMSL Research
Start Date
2007-06-19
End Date
2008-06-22
Status
Closed
Released Data Link
Team
Principal Investigator
Team Members
Related Publications
31st Annual Solid-State NMR meeting,Breckenridge, CO, July 2008.
poster presentation
Comparative Dynamics of Leucine Methyl Groups in FMOC-Leucine and in a Protein Hydrophobic Core Probed by Solid-State Deuteron Nuclear Magnetic Resonance over 7-324 K Temperature Range. Liliya Vugmeyster, Dmitry Ostrovsky, Mark Moses, Joseph J. Ford, Andrew S. Lipton, Gina L. Hoatson, and Robert L. Vold , J. Phys. Chem. B. Web Publication November 15, 2010; doi: 10.1021/jp1082467
Probing the Dynamics of a Protein Hydrophobic Core by Deuteron Solid-State Nuclear Magnetic Resonance Spectroscopy
Liliya Vugmeyster, Dmitry Ostrovsky, Joseph J. Ford, Sarah D. Burton, Andrew S. Lipton, Gina L. Hoatson, and Robert L. Vold
Publication Date (Web): September 3, 2009 | doi: 10.1021/ja902977u