The long-term objectives of this program project are to develop computational tools that can be used to extract accurate structural and dynamical information from site-directed spin-labeling (SDSL) studies on proteins. These new tools constitute an essential enabling technology to determine the structures and functional dynamics for a wide range of soluble and membrane-bound proteins and their complexes. As such, these tools should have an immediate and significant impact on SDSL studies of proteins here at Vanderbilt and elsewhere, and we intend to make all software and computational protocols developed in this program project freely available to colleagues at other institutions.
The specific aims of this program project are: 1) develop molecular dynamics and molecular modeling protocols to enable us to directly correlate EPR measurements of spin label mobility and accessibility with protein structure and structural fluctuations;2) develop simulation tools and protocols so that EPR measurements of inter-probe distances can be related directly to protein structure;3) develop a general computational protocol so that information obtained in Specific Aims 1 and 2 can be used to build and refine structural models;and 4) apply the tools and strategies developed in Aims 1-3 to SDSL data obtained for T4 lysozyme, a model protein of known structure, for CDB3, a more complex erythrocyte membrane protein that is to date only partially characterized structurally, and for amyloid precursor protein peptides that are not well characterized structurally at this time. These specific protein spectroscopic studies will provide the requisite experimental data necessary to develop and test the computational protocols. This program includes investigators with extensive eperience in EPR spectroscopy and SDSL studies of proteins (Beth and Hustedt), membrane protein structure-function studies (Beth, Lybrand, and Sanders), and use of molecular modeling and molecular simulation tools to refine three-dimensional protein structures from distance data obtained from spectroscopic and other biophysical studies (Lybrand and Smith).

National Institute of Health (NIH)
National Institute of General Medical Sciences (NIGMS)
Research Program Projects (P01)
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Special Emphasis Panel (ZRG1-BCMB-P (40))
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Wehrle, Janna P
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Vanderbilt University Medical Center
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Stein, Richard A; Beth, Albert H; Hustedt, Eric J (2015) A Straightforward Approach to the Analysis of Double Electron-Electron Resonance Data. Methods Enzymol 563:531-67
Edwards, Sarah J; Moth, Christopher W; Kim, Sunghoon et al. (2014) Automated structure refinement for a protein heterodimer complex using limited EPR spectroscopic data and a rigid-body docking algorithm: a three-dimensional model for an ankyrin-CDB3 complex. J Phys Chem B 118:4717-26
Smith, Jarrod A; Edwards, Sarah J; Moth, Christopher W et al. (2013) TagDock: an efficient rigid body docking algorithm for oligomeric protein complex model construction and experiment planning. Biochemistry 52:5577-84
Pester, Oxana; Barrett, Paul J; Hornburg, Daniel et al. (2013) The backbone dynamics of the amyloid precursor protein transmembrane helix provides a rationale for the sequential cleavage mechanism of γ-secretase. J Am Chem Soc 135:1317-29
Barrett, Paul J; Chen, Jiang; Cho, Min-Kyu et al. (2013) The quiet renaissance of protein nuclear magnetic resonance. Biochemistry 52:1303-20
Song, Yuanli; Hustedt, Eric J; Brandon, Suzanne et al. (2013) Competition between homodimerization and cholesterol binding to the C99 domain of the amyloid precursor protein. Biochemistry 52:5051-64
Zhuang, Tiandi; Chen, Qiuyan; Cho, Min-Kyu et al. (2013) Involvement of distinct arrestin-1 elements in binding to different functional forms of rhodopsin. Proc Natl Acad Sci U S A 110:942-7
Mittendorf, Kathleen F; Deatherage, Catherine L; Ohi, Melanie D et al. (2012) Tailoring of membrane proteins by alternative splicing of pre-mRNA. Biochemistry 51:5541-56
Deatherage, Catherine L; Hadziselimovic, Arina; Sanders, Charles R (2012) Purification and characterization of the human γ-secretase activating protein. Biochemistry 51:5153-9
Ghimire, Harishchandra; Hustedt, Eric J; Sahu, Indra D et al. (2012) Distance measurements on a dual-labeled TOAC AChR M2δ peptide in mechanically aligned DMPC bilayers via dipolar broadening CW-EPR spectroscopy. J Phys Chem B 116:3866-73

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