Characterization of Internal Protein Dynamics and Conformational Entropy by NMR Relaxation

Abstract

Recent studies suggest that the fast timescale motion of methyl-bearing side chains may play an important role in mediating protein activity. These motions have been shown to encapsulate the residual conformational entropy of the folded state that can potentially contribute to the energetics of protein function. Here, we provide an overview of how to characterize these motions using nuclear magnetic resonance (NMR) spin relaxation methods. The strengths and limitations of several techniques are highlighted in order to assist with experimental design. Particular emphasis is placed on the practical aspects of sample preparation, data collection, data fitting, and statistical analysis. Additionally, discussion of the recently refined “entropy meter” is presented and its use in converting NMR observables to conformational entropy is illustrated. Taken together, these methods should yield new insights into the complex interplay between structure and dynamics in protein function.

Protein conformational entropy is not slaved to water

“…Methyl and all aromatic order parameters are reported from a single sample. The precision of measured order parameters is generally better than a few percent 48 .…”

“…Model-free calculations were performed using in-house software 48 . Optimal tumbling models (isotropic, axially symmetric, or anisotropic) and rotational correlation times were determined 49 for each sample.…”

“…Order parameters and τ e values were determined using a grid search approach 50 , and errors were estimated using Monte Carlo. Details of these calculations are provided elsewhere 48 . Methyl O 2 axis values were derived from measured order parameters under the assumption of tetrahedral geometry (O 2 /0.111).…”

RING NMR dynamics: software for analysis of multiple NMR relaxation experiments

“…These two important attributes mean that NMR can probe dynamics with atomic spatial resolution and with a temporal resolution matched to the biological process in question. This ability to examine different atomic types and different timescales has led to the development of a wide range of NMR experiments, each optimized for a different dynamics phenomenon and different time scale (Sekhar and Kay 2019;Stetz et al 2019;Marušič et al 2019).…”

Dynamic 15N{1H} NOE measurements: a tool for studying protein dynamics

“…Since its first use of magnetic relaxation measurements of 15 N nuclei applied to the protein, the staphylococcal nuclease (Kay et al 1989), this method has become indispensable in the determination of molecular motions in biopolymers (Jarymowycz and Stone 2006;Kempf and Loria 2003;Palmer, III 2004;Reddy and Rayney 2010;Stetz et al 2019).…”