High-Resolution 1H Solid-State NMR Spectroscopy Using Windowed LG4 Homonuclear Dipolar Decoupling

We investigate the optimal implementation of windowed LG4 decoupling for the direct acquisition of high-resolution ¹H solid-state NMR spectra in the moderate magic-angle spinning regime (ω_r<2π × 35 kHz). We determine how the optimal value of the azimuth of the LG rotation axis, α, is affected by the choice of windowing parameters. We find that for both the windowless and windowed implementation of LG4, the optimal value of α is that for which the chemical-shift scaling factor is approximately 0.43. We then provide an analytical expression for estimating the chemical-shift scaling factor as a function of both α and the windowed decoupling parameters, which allows the optimal value of α to be easily determined for any implementation of LG4. We also introduce a simple and general method for improving the performance of any phase-modulated homonuclear dipolar decoupling sequence by compensating for pulse imperfection errors that accumulate in windowed spectra due to the repeated switching of the RF irradiation. This method involves the introduction of short (<0.5 µs) compensation pulses at the start and end of each decoupling pulse, the duration and phase of which are chosen to cancel the effects of the amplitude and phase transients that arise as a result of the RF switching. We demonstrate how this method can be implemented for homonuclear dipolar decoupling sequences such as wPMLG and wLG4 to reduce the chemical-shift scaling factor distortions generated by phase transients such that high-quality, high-resolution ¹H NMR spectra can be acquired at arbitrary RF offsets.     

RNA Dynamics by NMR Spectroscopy

An ever-increasing number of functional RNAs require a mechanistic understanding. RNA function relies on changes in its structure, so-called dynamics. To reveal dynamic processes and higher energy structures, new NMR methods have been developed to elucidate these dynamics in RNA with atomic resolution. In this Review, we provide an introduction to dynamics novices and an overview of methods that access most dynamic timescales, from picoseconds to hours. Examples are provided as well as insight into theory, data acquisition and analysis for these different methods. Using this broad spectrum of methodology, unprecedented detail and invisible structures have been obtained and are reviewed here. RNA, though often more complicated and therefore neglected, also provides a great system to study structural changes, as these RNA structural changes are more easily defined—Lego like—than in proteins, hence the numerous revelations of RNA excited states.     

A Comparative Study of Water- and Benzene-Saturated Mesoporous Silicas by 1H NMR Spectroscopy

1H NMR investigations have been carried out on water- and benzene-saturated mesoporous sillicas with a wide range of pore sizes (13–42 Å). The freezing point of the bound water increases with the pore size, approaching that of bulk water at large pore sizes. With large pore sizes, there is more than one transition due to the bound water species. The benzene-saturated samples show a behavior similar to that of water-saturated samples, although the freezing point depression is significantly greater. The pore size distribution generally becomes broader with the increase in pore size. The original Kelvin equation is satisfactory to estimate the pore radii both in the case of water and benzene-saturated samples.     

Applications of Two-Dimensional 1H NMR Methods to Photo-Chemically Induced Dynamic Nuclear Polarisation Spectroscopy

Photo-CIDNP is a useful technique to identify surface residues in proteins and to assign resonances in their ¹H NMR spectra by the spectral simplification that is obtained. In spite of this simplification, conventional photo-CIDNP spectra can be quite complex due to spectral overlap. For this reason two-dimensional (2D) methods are welcome. We present combinations of photo-CIDNP with 2D J-correlated spectroscopy (CIDNP-COSY) and with 2D NOE spectroscopy (CIDNP-NOESY). In these experiments we insert a saturation pulse sequence and a short laser-irradiation period at the beginning of the preparation period to generate CIDNP, leaving the remaining of the pulse scheme essentially unaltered. The CIDNP-COSY and CIDNP-NOESY spectra of a hen egg-white (HEW) lysozyme are presented. The CIDNP-NOESY spectrum of lysozyme is compared with the results from 1D cross-polarisation experiments on the same protein. CIDNP-NOESY spectra of lysozyme with and without inhibitor NAG are compared.     

Esterase Activity of Serum Albumin Studied by 1H NMR Spectroscopy and Molecular Modelling

Serum albumin possesses esterase and pseudo-esterase activities towards a number of endogenous and exogenous substrates, but the mechanism of interaction of various esters and other compounds with albumin is still unclear. In the present study, proton nuclear magnetic resonance (¹H NMR) has been applied to the study of true esterase activity of albumin, using the example of bovine serum albumin (BSA) and p-nitrophenyl acetate (NPA). The site of BSA esterase activity was then determined using molecular modelling methods. According to the data obtained, the accumulation of acetate in the presence of BSA in the reaction mixture is much more intense as compared with the spontaneous hydrolysis of NPA, which indicates true esterase activity of albumin towards NPA. Similar results were obtained for p-nitophenyl propionate (NPP) as substrate. The rate of acetate and propionate release confirms the assumption that there is a site of true esterase activity in the albumin molecule, which is different from the site of the pseudo-esterase activity Sudlow II. The results of molecular modelling of BSA and NPA interaction make it possible to postulate that Sudlow site I is the site of true esterase activity of albumin.