Amir Goldbourt

Sensitivity enhancement of the MQMAS NMR experiment by fast amplitude modulation of the pulses

Abstract We report here an improved way of doing the multiple-quantum magic-angle spinning (MQMAS) NMR experiment that relies on the use of amplitude modulated pulses. These pulses were found to yield MQMAS NMR signals that are considerably stronger (≈200–300%) than the ones arising from the usual continuous wave pulse schemes by virtue of a superior efficiency of the triple- to single-quantum conversion process. Numerical simulations and experimental results taking 23 N a and 87 R b nuclei as examples are presented that corroborate the usefulness of this approach.

Fast radio-frequency amplitude modulation in multiple-quantum magic-angle-spinning nuclear magnetic resonance: Theory and experiments

Multiple-quantum magic-angle-spinning (MQMAS NMR) spectroscopy has become a routine method to obtain high-resolution spectra of quadrupolar nuclei. One of the main problems in the performance of this experiment has been the poor efficiency of the radio-frequency pulses used in converting multiple-quantum coherences to the observable single-quantum signals. As the MQMAS experiment is basically an echo experiment this problem can be related to the efficiency with which continuous wave pulses can normally achieve the multiple- to single-quantum conversion for different crystallites in a spinning powdered sample. In this paper we investigate various aspects involved in this multiple-to-single quantum conversion, in the hope to facilitate the devise of new experimental schemes that can lead to significant MQMAS signal enhancements. We examine in particular a recently suggested experiment for MQMAS spectroscopy which employs amplitude-modulated radio-frequency pulses, and which can yield substantial signal and ...

Enhanced conversion of triple to single-quantum coherence in the triple-quantum MAS NMR spectrosocopy of spin-5/2 nuclei

Abstract A new type of fast amplitude modulated pulse scheme is presented here that yields a significant sensitivity enhancement in the triple-quantum magic angle spinning NMR spectrum of a spin-5/2 nucleus. Enhancement is achieved by fast phase alternation of the triple to single-quantum conversion pulse, which transfers triple to single-quantum coherence in a direct, non-adiabatic manner. The success of this pulse relies on the difference between the quadrupolar frequency and the intensity of the rf irradiation field, during its short segments. Numerical optimizations and experimental results are presented showing the improved performance.

The NMR-Rosetta capsid model of M13 bacteriophage reveals a quadrupled hydrophobic packing epitope.

Significance We present an atomic-resolution structure of the M13 filamentous bacteriophage capsid, one of many filamentous viruses that play important roles in many areas of research. The model was obtained by combining magic-angle spinning NMR and Rosetta modeling, used for the first time, to our knowledge, to derive the atomic structure of an intact virus capsid. The structure is made up of thousands of identical helical subunits stabilized by repeating hydrophobic pockets, which serve as a locking motif, suggesting a direct role in phage particle assembly. Analysis of various phage sequences suggests the presence of a conserved design principle for helical capsids. Because the current method does not rely on any particular preparation procedure, it can be applied to other viral capsids and molecular assemblies. Filamentous phage are elongated semiflexible ssDNA viruses that infect bacteria. The M13 phage, belonging to the family inoviridae, has a length of ∼1 μm and a diameter of ∼7 nm. Here we present a structural model for the capsid of intact M13 bacteriophage using Rosetta model building guided by structure restraints obtained from magic-angle spinning solid-state NMR experimental data. The C5 subunit symmetry observed in fiber diffraction studies was enforced during model building. The structure consists of stacked pentamers with largely alpha helical subunits containing an N-terminal type II β-turn; there is a rise of 16.6–16.7 Å and a tilt of 36.1–36.6° between consecutive pentamers. The packing of the subunits is stabilized by a repeating hydrophobic stacking pocket; each subunit participates in four pockets by contributing different hydrophobic residues, which are spread along the subunit sequence. Our study provides, to our knowledge, the first magic-angle spinning NMR structure of an intact filamentous virus capsid and further demonstrates the strength of this technique as a method of choice to study noncrystalline, high-molecular-weight molecular assemblies.

Multiple-Quantum Magic-Angle Spinning: High-Resolution Solid-State NMR of Half-Integer Spin Quadrupolar Nuclei

Abstract Of the several techniques introduced to obtain high-resolution solid-state NMR spectra of nuclei with half-integer spins larger than one-half, the multiple-quantum magic-angle-spinning (MQMAS) method remains the most popular. This technique is easy to implement and spectral analysis is straightforward. MQMAS has now become the method of choice for the characterization of materials containing half-integer spin quadrupolar nuclei such as 11 B, 17 O, 23 Na, and 27 Al. The combination of MQMAS with other solid-state NMR techniques has resulted in hetero- and homo-correlation experiments. They have helped in obtaining connectivity information and are expected to yield a wealth of geometry constraints in systems of quadrupolar nuclei. In this chapter, we review the technique of MQMAS in detail covering the basic theory, experimental implementation, signal optimization, data analysis and interpretation, signal enhancement schemes, applications, and give an overview of the hetero- and homo-correlation experiments involving MQMAS.

Intersubunit Hydrophobic Interactions in Pf1 Filamentous Phage

Magic angle spinning solid-state NMR has been used to study the structural changes in the Pf1 filamentous bacteriophage, which occur near 10 °C. Comparisons of NMR spectra recorded above and below 10 °C reveal reversible perturbations in many NMR chemical shifts, most of which are assigned to atoms of hydrophobic side chains of the 46-residue subunit. The changes mainly involve groups located in patches on the interfaces between neighboring capsid subunits. The observations show that the transition adjusts the hydrophobic interfaces between fairly rigid subunits. The low temperature form has been generally more amenable to structure determination; spin diffusion experiments on this form revealed unambiguous contacts between side chains of neighboring subunits. These contacts are important constraints for structure modeling.

Biomolecular magic-angle spinning solid-state NMR: recent methods and applications.

The link of structure and dynamics of biomolecules and their complexes to their function and to many cellular processes has driven the quest for their detailed characterization by a variety of biophysical techniques. Magic-angle spinning solid-state nuclear magnetic resonance spectroscopy provides detailed information on the structural properties of such systems and in particular contributes invaluable information on non-soluble, large molecular-weight and non-crystalline biomolecules. This review summarizes the recent progress that has been made in the characterization of macromolecular assemblies, viruses, membrane proteins, amyloid fibrils, protein aggregates and more by magic-angle spinning solid-state NMR.

Magic Angle Spinning NMR Spectroscopy: A Versatile Technique for Structural and Dynamic Analysis of Solid-Phase Systems

Magic Angle Spinning (MAS) NMR spectroscopy is a powerful method for analysis of a broad range of systems, including inorganic materials, pharmaceuticals, and biomacromolecules. The recent developments in MAS NMR instrumentation and methodologies opened new vistas to atomic-level characterization of a plethora of chemical environments previously inaccessible to analysis, with unprecedented sensitivity and resolution.

Determination of the lithium binding site in inositol monophosphatase, the putative target for lithium therapy, by magic-angle-spinning solid-state NMR.

Inositol monophosphatase (IMPase) catalyzes the hydrolysis of inositol monophosphate to inorganic phosphate and inositol. For this catalytic process to occur, Mg(2+) cations must exist in the active site. According to the inositol depletion hypothesis, IMPase activity is assumed to be higher than normal in patients suffering from bipolar disorder. Treatment with Li(+), an inhibitor of IMPase, reduces its activity, but the mechanism by which lithium exerts its therapeutic effects is still at a stage of conjecture. The Escherichia coli SuhB gene product possesses IMPase activity, which is also strongly inhibited by Li(+). It has significant sequence similarity to human IMPase and has most of its key active-site residues. Here we show that by using (7)Li magic-angle-spinning solid-state NMR spectroscopy, including {(13)C}(7)Li dipolar recoupling experiments, the bound form of lithium in the active site of wild-type E. coli SuhB can be unambiguously detected, and on the basis of our data and other biochemical data, lithium binds to site II, coupled to aspartate residues 84, 87, and 212.

Efficient rotational echo double resonance recoupling of a spin-1/2 and a quadrupolar spin at high spinning rates and weak irradiation fields.

A modification of the rotational echo (adiabatic passage) double resonance experiments, which allows recoupling of the dipolar interaction between a spin-1/2 and a half integer quadrupolar spin is proposed. We demonstrate efficient and uniform recoupling at high spinning rates (nu(r)), low radio-frequency (RF) irradiation fields (nu(1)), and high values of the quadrupolar interaction (nu(q)) that correspond to values of alpha=nu(1)(2)/nu(q)nu(r), the adiabaticity parameter, which are down to less than 10% of the traditional adiabaticity limit for a spin-5/2 (alpha=0.55). The low-alpha rotational echo double resonance curve is obtained when the pulse on the quadrupolar nucleus is extended to full two rotor periods and beyond. For protons (spin-1/2) and aluminum (spin-5/2) species in the zeolite SAPO-42, a dephasing curve, which is significantly better than the regular REAPDOR experiment (pulse length of one-third of the rotor period) is obtained for a spinning rate of 13 kHz and RF fields down to 10 and even 6 kHz. Under these conditions, alpha is estimated to be approximately 0.05 based on an average quadrupolar coupling in zeolites. Extensive simulations support our observations suggesting the method to be robust under a large range of experimental values.