This research is focused on the development of new techniques, based on the hydrophobicity of their surfaces, for separating, characterizing, and manipulating proteins. (The proposal is an extensively revised version of a previous proposal that was not funded.) Improving the methods used to separate, characterize, and manipulate proteins is broadly important in biochemistry and molecular biology: areas of application include proteomics and systems biology, purification of protein therapeutics, use in lab-on-a-chip analytical systems for clinical analyses, rational drug design, and fundamental protein science. The proposed research has five specific objectives: 1. To develop a new, validated model system for studying the biophysics of hydrophobic regions of protein surfaces. 2. To develop new techniques for characterization of proteins based on their susceptibility to denaturation by hydrophobic surfactants and hydrotropes (e.g., SDS, not urea). 3. To rationalize systems that use hydrophobic surfactants in manipulating proteins. 4. To provide a solution to the problem of adsorption of proteins on the hydrophobic walls of microchannels in plastic LoC systems. 5. To help to understand the influence of hydrophobicity and organic character of the medium on binding of ligands to proteins. Lay Description: This research will contribute to the analysis and purification of proteins used in treatment of human disease, and to the development of instrumental techniques useful in the diagnosis of disease, in screening for public health, and in response to biological events (emerging disease, bioterrorism).

National Institute of Health (NIH)
National Institute of General Medical Sciences (NIGMS)
Research Project (R01)
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Enabling Bioanalytical and Biophysical Technologies Study Section (EBT)
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Edmonds, Charles G
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Harvard University
Schools of Arts and Sciences
United States
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Shaw, Bryan F; Schneider, Gregory F; Whitesides, George M (2012) Effect of surfactant hydrophobicity on the pathway for unfolding of ubiquitin. J Am Chem Soc 134:18739-45
Mecinovic, Jasmin; Snyder, Phillip W; Mirica, Katherine A et al. (2011) Fluoroalkyl and alkyl chains have similar hydrophobicities in binding to the ""hydrophobic wall"" of carbonic anhydrase. J Am Chem Soc 133:14017-26
Snyder, Phillip W; Mecinovic, Jasmin; Moustakas, Demetri T et al. (2011) Mechanism of the hydrophobic effect in the biomolecular recognition of arylsulfonamides by carbonic anhydrase. Proc Natl Acad Sci U S A 108:17889-94
Shaw, Bryan F; Schneider, Gregory F; Arthanari, Haribabu et al. (2011) Complexes of native ubiquitin and dodecyl sulfate illustrate the nature of hydrophobic and electrostatic interactions in the binding of proteins and surfactants. J Am Chem Soc 133:17681-95
Lee, Andrew; Mirica, Katherine A; Whitesides, George M (2011) Influence of fluorocarbon and hydrocarbon acyl groups at the surface of bovine carbonic anhydrase II on the kinetics of denaturation by sodium dodecyl sulfate. J Phys Chem B 115:1199-210
Lee, Andrew; Tang, Sindy K Y; Mace, Charles R et al. (2011) Denaturation of proteins by SDS and tetraalkylammonium dodecyl sulfates. Langmuir 27:11560-74
Lee, Andrew; Whitesides, George M (2010) Analysis of inorganic polyphosphates by capillary gel electrophoresis. Anal Chem 82:6838-46
Shaw, Bryan F; Arthanari, Haribabu; Narovlyansky, Max et al. (2010) Neutralizing positive charges at the surface of a protein lowers its rate of amide hydrogen exchange without altering its structure or increasing its thermostability. J Am Chem Soc 132:17411-25
Krishnamurthy, Vijay M; Kaufman, George K; Urbach, Adam R et al. (2008) Carbonic anhydrase as a model for biophysical and physical-organic studies of proteins and protein-ligand binding. Chem Rev 108:946-1051
Schneider, Gregory F; Shaw, Bryan F; Lee, Andrew et al. (2008) Pathway for unfolding of ubiquitin in sodium dodecyl sulfate, studied by capillary electrophoresis. J Am Chem Soc 130:17384-93

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