Optimizing ssNMR experiments for dilute proteins in heterogeneous mixtures at high magnetic fields

Abstract

Solid-state NMR spectroscopy at high magnetic fields is proving to be an effective technique in structural biology, particularly for proteins which are not amenable to traditional X-ray and solution NMR approaches. Several parameters can be selected to provide optimal sensitivity, improve sample stability, and ensure biological relevance for ssNMR measurements on protein samples. These include selection of sample conditions, NMR probe design, and design of pulse experiments. Here, we demonstrate and evaluate several engineering and experimental approaches for pursuing measurements on dilute proteins in heterogeneous mixtures.

1H-detected solid-state NMR of proteins entrapped in bioinspired silica: a new tool for biomaterials characterization

“…Another important point is the amount of measurement time that is needed: a 1 H-start/ 1 H detect experiment is expected to be roughly a factor 2.5 more sensitive than a 1 H-start- 13 C detect experiment, neglecting relaxation effects and assuming 100% efficiency in each transfer step; the amount of experiments to be acquired in the indirect detection is about a factor 6 smaller and the full-rotor sensitivity of a solenoid coil (including the higher efficiency but also the reduction of sample) on passing from a 4 mm to a 1.3 mm rotor is roughly reduced by a factor 9.5 5150, which would yield about a factor 2 increase in sensitivity, corresponding to about a factor 4 reduction in experiment time, thus in experiment price. These consideration are qualitatively reflected in the present case, as we find that the 1 H- 15 N HSQC experiment could be acquired in 10 h 36′ for catMMP12-R5 in the current setup, whereas the 13 C- 13 C DARR experiment reported in reference41 was acquired in 45 h 31′.…”

Low-E probe for 19F–1H NMR of dilute biological solids

“…Finally, the elimination of fluorinated materials from the probe, which involved modification of trimmer capacitors and construction of custom 50 X coaxial cables, resulted in a 1 MHz-wide spectral window free of 19 F background NMR signal after 10 4 averages. The low-E 19 F-1 H probehead design described in this paper need not be limited to the static applications and could be implemented in MAS stators in a manner similar to that discussed in [28].…”

Multivariate Curve Resolution for 2D Solid-State NMR spectra

“…However, not only are high field magnets more expensive than low field magnets because of the different manufacturing processes, but their operational costs are also higher, so that, overall, the price per experiment gets significantly higher even when moving from 16.4 to 18.8 T (from 700 to 800 MHz). Another option is to use larger amounts of sample, but (a) the amount of sample is likely limited, (b) small rotors need to be used to increase the maximal achievable spinning speed and thus the resolution, and (c) more components may be present, imposing a strong dilution on the species of interest. , Proton detection, available at high spinning frequency, can be used to increase the sensitivity, ,, but this is necessarily limited to proton-rich materials. Dynamic nuclear polarization (DNP) is also a viable route to study silicon-based materials, − but the equipment is more expensive than standard NMR.…”