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Solvation in protein (un)folding of melittin tetramer-monomer transition.

ABSTRACT: Protein structural integrity and flexibility are intimately tied to solvation. Here, we examine the effect that changes in bulk and local solvent properties have on protein structure and stability. We observe the change in solvation of an unfolding of the protein model, melittin, in the presence of a denaturant, trifluoroethanol. The peptide system displays a well defined transition in that the tetramer unfolds without disrupting the secondary or tertiary structure. In the absence of local structural perturbation, we are able to reveal exclusively the role of solvation dynamics in protein structure stabilization and the (un)folding pathway. A sudden retardation in solvent dynamics, which is coupled to the change in protein structure, is observed at a critical trifluoroethanol concentration. The large amplitude conformational changes are regulated by the local solvent hydrophobicity and bulk solvent viscosity.

SUBMITTER: Othon CM

PROVIDER: S-EPMC2722364 | biostudies-literature | 2009 Aug

REPOSITORIES: biostudies-literature

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Solvation in protein (un)folding of melittin tetramer-monomer transition.

Othon Christina M CM Kwon Oh-Hoon OH Lin Milo M MM Zewail Ahmed H AH

Proceedings of the National Academy of Sciences of the United States of America 20090721 31

Protein structural integrity and flexibility are intimately tied to solvation. Here, we examine the effect that changes in bulk and local solvent properties have on protein structure and stability. We observe the change in solvation of an unfolding of the protein model, melittin, in the presence of a denaturant, trifluoroethanol. The peptide system displays a well defined transition in that the tetramer unfolds without disrupting the secondary or tertiary structure. In the absence of local struc ...[more]

PMID: 19622745

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Project description:Ficoll, an inert macromolecule, is a common in vitro crowder, but by itself it does not reproduce in-cell stability or kinetic trends for protein folding. Lysis buffer, which contains ions, glycerol as a simple kosmotrope, and mimics small crowders with hydrophilic/hydrophobic patches, can reproduce sticking trends observed in cells but not the crowding. We previously suggested that the proper combination of Ficoll and lysis buffer could reproduce the opposite in-cell folding stability trend of two proteins: variable major protein-like sequence expressed (VlsE) is destabilized in eukaryotic cells and phosphoglycerate kinase (PGK) is stabilized. Here, to discover a well-characterized solvation environment that mimics in-cell stabilities for these two very differently behaved proteins, we conduct a two-dimensional scan of Ficoll (0-250 mg/ml) and lysis buffer (0-75%) mixtures. Contrary to our previous expectation, we show that mixtures of Ficoll and lysis buffer have a significant nonadditive effect on the folding stability. Lysis buffer enhances the stabilizing effect of Ficoll on PGK and inhibits the stabilizing effect of Ficoll on VlsE. We demonstrate that a combination of 150 mg/ml Ficoll and 60% lysis buffer can be used as an in vitro mimic to account for both crowding and non-steric effects on PGK and VlsE stability and folding kinetics in the cell. Our results also suggest that this mixture is close to the point where phase separation will occur. The simple mixture proposed here, based on commercially available reagents, could be a useful tool to study a variety of cytoplasmic protein interactions, such as folding, binding and assembly, and enzymatic reactions. SIGNIFICANCE STATEMENT: The complexity of the in-cell environment is difficult to reproduce in the test tube. Here we validate a mimic of cellular crowding and sticking interactions in a test tube using two proteins that are differently impacted by the cell: one is stabilized and the other is destabilized. This mimic is a starting point to reproduce cellular effects on a variety of protein and biomolecular interactions, such as folding and binding.

Dataset Information

Solvation in protein (un)folding of melittin tetramer-monomer transition.

Publications

Solvation in protein (un)folding of melittin tetramer-monomer transition.

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