On the Use of Side‐Chain NMR Relaxation Data to Derive Structural and Dynamical Information on Proteins: A Case Study Using Hen Lysozyme

Abstract

Values of S2CH and S2NH order parameters derived from NMR relaxation measurements on proteins cannot be used straightforwardly to determine protein structure because they cannot be related to a single protein structure, but are defined in terms of an average over a conformational ensemble. Molecular dynamics simulation can generate a conformational ensemble and thus can be used to restrain S2CH and S2NH order parameters towards experimentally derived target values S2CH (exp) and S2NH (exp). Application of S2CH and S2NH order‐parameter restraining MD simulation to bond vectors in 63 side chains of the protein hen egg white lysozyme using 51 S2CH (exp) target values and 28 S2NH (exp) target values shows that a conformational ensemble compatible with the experimentally derived data can be obtained by using this technique. It is observed that S2CH order‐parameter restraining of C−H bonds in methyl groups is less reliable than S2NH order‐parameter restraining because of the possibly less valid assumptions and approximations used to derive experimental S2CH (exp) values from NMR relaxation measurements and the necessity to adopt the assumption of uniform rotational motion of methyl C−H bonds around their symmetry axis and of the independence of these motions from each other. The restrained simulations demonstrate that side chains on the protein surface are highly dynamic. Any hydrogen bonds they form and that appear in any of four different crystal structures, are fluctuating with short lifetimes in solution.

The ‘Shape-Shifter’ Peptide from the Disulphide Isomerase PmScsC Shows Context-Dependent Conformational Preferences

“…Under solution conditions, any hydrogen bonds between molecules in different trimers will clearly be missing. In addition, any intramolecular or inter-monomer hydrogen bonds involving side chain groups on the surface of the protein are expected to be fluctuating with short lifetimes [ 30 ]. Consequently, the quaternary structure of the trimer is likely to be an ensemble of conformers, rather than any one of the different structures seen in crystals predominating in solution.…”

A Suite of Advanced Tutorials for the GROMOS Biomolecular Simulation Software [Article v1.0]

“…Moreover, a possible force-field deficiency hampering the agreement with experiment can be redressed using this restraining technique. In this way configurational ensembles consistent with NMR data can be generated allowing a structural interpretation of experimental observations [14,15]. We will demonstrate the use of time-averaged order parameters by means of the third IgG-binding domain of Protein G (GB3), which is a small 56-residue protein.…”

On the use of 3J-coupling NMR data to derive structural information on proteins

“…of r XY is essentially constant over time and thus equal to its effective value r eff XY . The set of 79 experimentally derived S 2 values (Buck et al 1995;Moorman et al 2012) together with the S 2 values calculated from the five MD simulations started from the 2VB1 X-ray crystal structure (Smith et al 2020) can be found in Supporting Information, Table S2.…”

“…Before calculating S 2 XY , the protein trajectory structures are superimposed using the backbone atoms (N, C α , C) of residues 3-126 in the fit in order to eliminate the effect of overall rotation of the protein upon the S 2 XY -values. Use of only the backbone atoms of four of the five α-helices and two β-strands in [24][25][26][27][28][29][30][31][32][33][34][35][36][41][42][43][44][45][50][51][52][53][89][90][91][92][93][94][95][96][97][98][99][108][109][110][111][112][113][114][115] CD2 -values for Leu residues (Smith et al 2020).…”

“…1-2 2-3 3-4 4-5 > 5 (Smith et al 2020), in the four unrestrained MD simulations starting from four different X-ray crystal structures and in the 3 J-coupling time-averaging local-elevation restrained MD simulations starting from the 2VB1 X-ray crystal structure and using different sets of backbone and side-chain restraints. Order parameter values are reported in Table S2 in Supporting Information…”