[NMR paper] Probing the Residual Structure of the Low Populated Denatured State of ADA2h under Folding Conditions by Relaxation Dispersion NMR Spectroscopy.

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Probing the Residual Structure of the Low Populated Denatured State of ADA2h under Folding Conditions by Relaxation Dispersion NMR Spectroscopy.

Related Articles Probing the Residual Structure of the Low Populated Denatured State of ADA2h under Folding Conditions by Relaxation Dispersion NMR Spectroscopy.

Biochemistry. 2015 Jun 26;

Authors: Pustovalova Y, Kukic P, Vendruscolo M, Korzhnev DM

Abstract
The structural characterization of low-populated states of proteins with accuracy comparable to that achievable for native states is important for understanding the mechanisms of protein folding and function, as well as misfolding and aggregation. Because of the transient nature of these low-populated states, they are seldom detected directly under conditions that favor folding. The activation domain of human procarboxypeptidase A2 (ADA2h) is an ?/?-protein that forms amyloid fibrils at low pH presumably initiated from a denatured state with a considerable amount of residual structure. Here we used Carr-Parcell-Meiboom-Gill relaxation dispersion (CPMG RD) NMR spectroscopy to characterize the structure of the denatured state of the ADA2h I71V mutant under conditions that favor folding. Under these conditions, the lifetime of the denatured state of the I71V ADA2h is of the order of milliseconds and its population is about several percent, which makes this mutant amenable to studies by CPMG RD methods. The nearly complete set of CPMG RD derived backbone 15N, 13C and 1H NMR chemical shifts in the I71V ADA2h denatured state reveals that it retains a significant fraction (up to 50-60%) of native-like ?-helical structure, while the regions encompassing native ?-strands are structured to a much lesser extent. The native-like ?-helical structure of the denatured state can bring together hydrophobic residues on the same sides of ?-helices, making them available for intra- or inter-molecular interactions. CPMG RD data analysis thus allowed a detailed structural characterization of the ADA2h denatured state under folding conditions not previously achieved for this protein.


PMID: 26115097 [PubMed - as supplied by publisher]



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