This project involves development of a new method, Multi-wavelength Anomalous Diffraction using Medium Angle X-ray solution scattering (MADMAX), for the precise measurement of interatomic distances within proteins and other macromolecules in aqueous solution. It will make possible the characterization of structural changes and intra-molecular movements that cannot be studied by existing techniques. Proteins are dynamic molecules whose activities contribute to all biological processes. Virtually all protein function requires intra- molecular movement, whether for the application of mechanical force, chemical transformation or molecular translocation. MADMAX will make possible accurate atomic-level measurement of these movements within proteins in solution. MADMAX should be generally applicable to macromolecules in solution and capable of elucidating protein action during a broad range of phenomena including allosteric interactions;protein-ligand interactions;channel gating;domain movements;and protein folding. It should be capable of measuring changes in the length of interatomic vectors with an accuracy of up to ~1 ?. It can be adapted for time- resolved studies - to at least millisecond resolution. The goals of this project are: (i) Demonstrate the accurate measurement of anomalous differences in solution scattering from well characterized proteins. (ii) Predict the anomalous differences from atomic coordinate sets. (iii) Determine the practical range of the method: How large a protein is this method applicable to? How many labels are required for detection? How much disorder in the label position can be tolerated? (iv) Demonstrate the use of MADMAX for precise measurement of structural changes due to ligand binding. (v) Demonstrate the use of MADMAX for the study of structural changes in membrane proteins. (vi) Demonstrate the use of time-resolved MADMAX for study of protein conformational changes. Taking full advantage of the capabilities of bacterial expression systems and chemical synthesis of proteins for the introduction of labels, the range of experiments that will be possible is almost limitless. Development of its full range of capabilities will establish MADMAX as an important new approach to the observation of protein structure and function in aqueous solution. This project will result in the development of a novel tool for the precise measurement of interatomic distances within proteins and the motions that they undergo during action. Development of advanced therapies depends on a deep understanding of these molecular actions because they form the basis of virtually all biological functions. Application of this approach to molecular systems involved in pathogenesis will contribute significantly to the development of strategies for the diagnosis and treatment of a broad range of human diseases.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM085648-03
Application #
7922550
Study Section
Special Emphasis Panel (ZGM1-GDB-7 (EU))
Program Officer
Wehrle, Janna P
Project Start
2008-08-01
Project End
2010-09-28
Budget Start
2010-08-01
Budget End
2010-09-28
Support Year
3
Fiscal Year
2010
Total Cost
$27,630
Indirect Cost
Name
University of Chicago
Department
Type
Organized Research Units
DUNS #
005421136
City
Chicago
State
IL
Country
United States
Zip Code
60637
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Onuk, A Emre; Akcakaya, Murat; Bardhan, Jaydeep P et al. (2015) Constrained Maximum Likelihood Estimation of Relative Abundances of Protein Conformation in a Heterogeneous Mixture from Small Angle X-Ray Scattering Intensity Measurements. IEEE Trans Signal Process 63:5383-5394
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Virtanen, J J; Sosnick, T R; Freed, K F (2014) Ionic strength independence of charge distributions in solvation of biomolecules. J Chem Phys 141:22D503
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Makowski, L; Bardhan, J; Gore, D et al. (2012) Multi-wavelength anomalous diffraction using medium-angle X-ray solution scattering (MADMAX). Biophys J 102:927-33
Bardhan, Jaydeep P; Jungwirth, Pavel; Makowski, Lee (2012) Affine-response model of molecular solvation of ions: Accurate predictions of asymmetric charging free energies. J Chem Phys 137:124101
Makowski, Lee; Gore, David; Mandava, Suneeta et al. (2011) X-ray solution scattering studies of the structural diversity intrinsic to protein ensembles. Biopolymers 95:531-42
Virtanen, Jouko Juhani; Makowski, Lee; Sosnick, Tobin R et al. (2011) Modeling the hydration layer around proteins: applications to small- and wide-angle x-ray scattering. Biophys J 101:2061-9

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