Site‐Resolved Backbone and Side‐Chain Intermediate Dynamics in a Carbohydrate‐Binding Module Protein Studied by Magic‐Angle Spinning NMR Spectroscopy

Abstract

Magic-angle spinning solid-state NMR spectroscopy has been applied to study the dynamics of CBM3b-Cbh9A from Clostridium thermocellum (ctCBM3b), a cellulose binding module protein. This 146-residue protein has a nine-stranded β-sandwich fold, in which 35 % of the residues are in the β-sheet and the remainder are composed of loops and turns. Dynamically averaged (1) H-(13) C dipolar coupling order parameters were extracted in a site-specific manner by using a pseudo-three-dimensional constant-time recoupled separated-local-field experiment (dipolar-chemical shift correlation experiment; DIPSHIFT). The backbone-Cα and Cβ order parameters indicate that the majority of the protein, including turns, is rigid despite having a high content of loops; this suggests that restricted motions of the turns stabilize the loops and create a rigid structure. Water molecules, located in the crystalline interface between protein units, induce an increased dynamics of the interface residues thereby lubricating crystal water-mediated contacts, whereas other crystal contacts remain rigid.

High resolution observed in 800 MHz DNP spectra of extremely rigid type III secretion needles

“…It has been shown that ssNMR is a useful tool for the investigation of protein dynamics (Ivanir-Dabora et al 2015;Lamley et al 2015;Lewandowski et al 2015;Ma et al 2015). In our previous study (Fricke et al 2014), we found that the peaks of the residues located on the outer surface of the needle are more difficult to identify than those that face towards the needle lumen.…”

Unifying Charge Generation, Recombination, and Extraction in Low‐Offset Non‐Fullerene Acceptor Organic Solar Cells

“…[94,[97][98][99] To distinguish between the two cases, we performed 1 H- 13 C DIPSHIFT experiments on both blends. [100][101][102] The objective of the experiment was to quantify the degree of motional averaging by measuring the apparent strength of the 1 H- 13 C dipolar couplings of CH or CH 2 moieties and comparing these to theoretical values. [101,103] Details of the experiment are provided in the Supporting Information; data obtained for both blends show no significant differences in the extents of motional averaging between the two.…”

Distance measurements to quadrupolar nuclei: Evolution of the rotational echo double resonance technique

“…Many elements that are of interest share close Larmor frequencies. For example, 59 Co, 69,71 Ga, 45 Sc, 55 Mn, 79,81 Br, 27 Al, 51 V, and 23 Na are all within 8% of the 13 C Larmor frequency. Also, 35 Cl/ 15 N, 133 Cs/ 17 O, 6 Li/ 2 H, and 7 Li/ 31 P are just some examples of spin pairs with similarly close Larmor frequencies.…”

“…(ii) Instead of fixing the pulse position at εT R , the value of ε itself is varied. This is the dipolar chemical shift correlation (DIPSHIFT) experiment, [22] which is highly useful for measuring protein dynamics (e.g., in previous studies [23][24][25] ). The similarity to REDOR has been extensively discussed.…”