Protein folding and aggregation are processes that constantly interact, sometimes leading to disease. Both protein folding and protein aggregation are sensitive to pressure, a thermodynamically simpler variable than temperature or solvent additives. We develop a new pressure jump simultaneously capable of >2500 atm pressure drops in <700 nanoseconds on <1 nM of protein sample. The experiment will be used to study the interplay of folding and aggregation of the Syrian hamster prion ShPrP, which is known to have very fast pressure-jump kinetic phases that could not be resolved by conventional instruments. Pressure- and concentration-dependent studies will reveal how formation of early folding intermediates and subsequent formation of the native state compete with transient aggregation that could eventually lead to formation of protofibrils. As a second pressure jump project, we also study in detail the folding of lambda repressor mutants designed to alter the packing and secondary structure propensity of the protein. For folding studies, the pressure drop provides an alternative refolding method that perturbs secondary structure less than temperature jump experiments. We picked lambda repressor because rich T-jump data already exist, and T-jump and P-jump experiments will be interesting to compare. All experiments will be analyzed with a folding or folding/aggregation master equation kinetics model that is useful for comparison with Markov dynamics derived from molecular dynamics simulation. We collaborate with the Schulten and Pande group who will carry out relevant simulations. The goal is to build a good low-resolution model of the folding or folding/aggregation energy landscape of these proteins.

Public Health Relevance

Many diseases are caused by the competition between folding and aggregation of proteins. Very fast and large pressure jumps are a new way of studying the interplay between folding and the earliest aggregation step of a model of the Creutzfeldt-Jakob disease protein.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM093318-03
Application #
8269853
Study Section
Special Emphasis Panel (ZRG1-BCMB-B (03))
Program Officer
Wehrle, Janna P
Project Start
2010-05-01
Project End
2013-08-31
Budget Start
2012-05-01
Budget End
2013-08-31
Support Year
3
Fiscal Year
2012
Total Cost
$260,175
Indirect Cost
$86,925
Name
University of Illinois Urbana-Champaign
Department
Chemistry
Type
Schools of Arts and Sciences
DUNS #
041544081
City
Champaign
State
IL
Country
United States
Zip Code
61820
Dave, Kapil; Jäger, Marcus; Nguyen, Houbi et al. (2016) High-Resolution Mapping of the Folding Transition State of a WW Domain. J Mol Biol 428:1617-36
Guin, Drishti; Sye, Kori; Dave, Kapil et al. (2016) Dodine as a transparent protein denaturant for circular dichroism and infrared studies. Protein Sci 25:1061-8
Dave, Kapil; Davtyan, Aram; Papoian, Garegin A et al. (2016) Environmental Fluctuations and Stochastic Resonance in Protein Folding. Chemphyschem 17:1341-8
Gelman, Hannah; Wirth, Anna Jean; Gruebele, Martin (2016) ReAsH as a Quantitative Probe of In-Cell Protein Dynamics. Biochemistry 55:1968-76
Zhang, Yi; Schulten, Klaus; Gruebele, Martin et al. (2016) Disulfide Bridges: Bringing Together Frustrated Structure in a Bioactive Peptide. Biophys J 110:1744-52
Dave, Kapil; Gelman, Hannah; Thu, Chu Thi Hien et al. (2016) The Effect of Fluorescent Protein Tags on Phosphoglycerate Kinase Stability Is Nonadditive. J Phys Chem B 120:2878-85
Prigozhin, Maxim B; Chao, Shu-Han; Sukenik, Shahar et al. (2015) Mapping fast protein folding with multiple-site fluorescent probes. Proc Natl Acad Sci U S A 112:7966-71
Wirth, Anna Jean; Liu, Yanxin; Prigozhin, Maxim B et al. (2015) Comparing Fast Pressure Jump and Temperature Jump Protein Folding Experiments and Simulations. J Am Chem Soc 137:7152-9
Dave, Kapil; Gruebele, Martin (2015) Fast-folding proteins under stress. Cell Mol Life Sci 72:4273-85
Goodman, J S; Chao, S-H; Pogorelov, T V et al. (2014) Filling up the heme pocket stabilizes apomyoglobin and speeds up its folding. J Phys Chem B 118:6511-8

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