[NMR paper] 19F NMR Studies of a Desolvated Near-Native Protein Folding Intermediate.

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19F NMR Studies of a Desolvated Near-Native Protein Folding Intermediate.

19F NMR Studies of a Desolvated Near-Native Protein Folding Intermediate.

Biochemistry. 2013 Aug 1;

Authors: Kitevski-Leblanc JL, Hoang J, Thach W, Larda ST, Prosser RS

Abstract
While many proteins are recognized to undergo folding via an intermediate, the microscopic nature of folding intermediates is less understood. In this study, 19F NMR and near UV circular dichroism (CD) are used to characterize a transition to a thermal folding intermediate of calmodulin, a water-soluble protein, which is biosynthetically enriched with 3-fluorophenylalanine (3F-Phe). 19F NMR solvent isotope shifts, resulting from replacing H2O with D2O, and paramagnetic shifts arising from dissolved O2 are used to monitor changes in water accessibility and hydrophobicity of the protein interior, as the protein progresses from a native state to an unfolded state, along a heat denaturation pathway. In comparison to the native state, solvent isotope shifts reveal the decreased presence of water in the hydrophobic core while paramagnetic shifts show increased hydrophobicity of this folding intermediate. 15N,1H and methyl 13C,1H HSQC NMR spectra identify that this folding intermediate retains a near-native tertiary structure, whose hydrophobic interior is highly dynamic. 19F NMR CPMG relaxation dispersion measurements suggest the near-native state is transiently adopted well below the temperature associated with its onset.


PMID: 23906334 [PubMed - as supplied by publisher]



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