This proposal is focused on studying side-chain and backbone motional dynamics in Streptococcal protein G B1 domain by using 13/C and 15/N-NMR relaxation and rotational model analyses. GB1 has only 56 amino acid residues arranged in a fold with a central alpha-helix packed against a four-stranded beta-sheet. Despite its small size, the thermodynamic properties are similar to those observed in larger proteins. GB1 lends itself well to selective 13/C/15/N isotropic enrichment via peptide synthesis and to experimental study using both NMR relaxation-derived auto-and cross-peptide synthesis and to experimental study using both NMR relation-derived auto-and cross-peptide synthesis and to experimental study using both NMR relation-derived auto- and cross-correlation spectral densities for analysis of all backbone and side-chain N-H and C-H motional vectors.
The specific aims of this project are: (1) to determine internal motions of all side-chain and backbone positions in unfolded states of GB1, and (3) to develop novel approaches and motional models to analyzes these NMR relaxation data. Most NMR dynamics studies reported to data, while qualitatively informative, are limited to analysis of auto-correlation parameters [usually from a single motional vector in a residue, i.e., backbone 15/N-H or 13/C/alpha-H] derived by using some model free approach. This grant stands apart from previous studies in that it will use both auto- and cross-correlation parameters from all available motional vectors (backbone and side-chain) and various rotational models and model free approaches to provide a more detailed description of motional dynamics in a protein. Dipolar and dipolar-CSA cross-correlation spectral densities, T/1, T/1p, T/2 and heteronuclear NOEs at at least two frequencies will be determined. For backbone motions, 13/C/alpha/H, 13/CO and 15/NH relaxation rates will be measured, while for side-chain motions, 13CH n auto- and cross- correlations will provide unique motional information.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM058005-02
Application #
6019468
Study Section
Molecular and Cellular Biophysics Study Section (BBCA)
Project Start
1998-08-01
Project End
2002-07-31
Budget Start
1999-08-01
Budget End
2000-07-31
Support Year
2
Fiscal Year
1999
Total Cost
Indirect Cost
Name
University of Minnesota Twin Cities
Department
Biochemistry
Type
Schools of Medicine
DUNS #
168559177
City
Minneapolis
State
MN
Country
United States
Zip Code
55455
Idiyatullin, Djaudat; Daragan, Vladimir A; Mayo, Kevin H (2004) A simple method to measure 13CH2 heteronuclear dipolar cross-correlation spectral densities. J Magn Reson 171:4-9
Idiyatullin, Djaudat; Nesmelova, Irina; Daragan, Vladimir A et al. (2003) Comparison of (13)C(alpha)H and (15)NH backbone dynamics in protein GB1. Protein Sci 12:914-22
Idiyatullin, Djaudat; Nesmelova, Irina; Daragan, Vladimir A et al. (2003) Heat capacities and a snapshot of the energy landscape in protein GB1 from the pre-denaturation temperature dependence of backbone NH nanosecond fluctuations. J Mol Biol 325:149-62
Idiyatullin, D; Daragan, V A; Mayo, K H (2001) Improved measurement of (15)N-[(1)H] NOEs in the presence of H(N)-water proton chemical exchange. J Magn Reson 153:138-43
Idiyatullin, D; Daragan, V A; Mayo, K H (2001) A new approach to visualizing spectral density functions and deriving motional correlation time distributions: applications to an alpha-helix-forming peptide and to a well-folded protein. J Magn Reson 152:132-48
Mayo, K H; Daragan, V A; Idiyatullin, D et al. (2000) Peptide internal motions on nanosecond time scale derived from direct fitting of (13)C and (15)N NMR spectral density functions. J Magn Reson 146:188-95