Varian, Inc.

About: Varian, Inc. is a based out in . It is known for research contribution in the topics: Electromagnetic coil & Ion. The organization has 337 authors who have published 380 publications receiving 8661 citations.

Proton-Detected Solid-State NMR Spectroscopy of Fully Protonated Proteins at 40 kHz Magic-Angle Spinning

Abstract: Remarkable progress in solid-state NMR has enabled complete structure determination of uniformly labeled proteins in the size range of 5-10 kDa. Expanding these applications to larger or mass-limited systems requires further improvements in spectral sensitivity, for which inverse detection of 13C and 15N signals with 1H is one promising approach. Proton detection has previously been demonstrated to offer sensitivity benefits in the limit of sparse protonation or with approximately 30 kHz magic-angle spinning (MAS). Here we focus on experimental schemes for proteins with approximately 100% protonation. Full protonation simplifies sample preparation and permits more complete chemical shift information to be obtained from a single sample. We demonstrate experimental schemes using the fully protonated, uniformly 13C,15N-labeled protein GB1 at 40 kHz MAS rate with 1.6-mm rotors. At 500 MHz proton frequency, 1-ppm proton line widths were observed (500 +/- 150 Hz), and the sensitivity was enhanced by 3 and 4 times, respectively, versus direct 13C and 15N detection. The enhanced sensitivity enabled a family of 3D experiments for spectral assignment to be performed in a time-efficient manner with less than a micromole of protein. CANH, CONH, and NCAH 3D spectra provided sufficient resolution and sensitivity to make full backbone and partial side-chain proton assignments. At 750 MHz proton frequency and 40 kHz MAS rate, proton line widths improve further in an absolute sense (360 +/- 115 Hz). Sensitivity and resolution increase in a better than linear manner with increasing magnetic field, resulting in 14 times greater sensitivity for 1H detection relative to that of 15N detection.

Solid‐State Protein‐Structure Determination with Proton‐Detected Triple‐Resonance 3D Magic‐Angle‐Spinning NMR Spectroscopy

Abstract: Advances over the last decade in magic-angle spinning solid-state NMR (MAS SSNMR) have enabled the complete structure determination of several small proteins.[1] In principle, SSNMR is not limited by molecular size, which is one major advantage over solution NMR in challenging applications such as membrane protein complexes and high molecular weight protein aggregates. However, solid-state structure determination of larger proteins is typically hindered by the low sensitivity and relatively short measurable distances imposed by the observation of nuclei with low gyromagnetic ratios (γ), such as 13C and 15N. The large γ of 1H, while providing high detection sensitivity and NOE distance restraints for solution NMR,[2] results in large dipolar couplings in the solid state, which may degrade both spectral resolution and sensitivity.[3] Recent studies by Reif and Zilm and their respective coworkers have demonstrated that these challenges in resolution and sensitivity can be overcome by using spin dilution, replacing all non-exchangeable protons with deuterons.[3] In combination with high magnetic fields and ~20 kHz MAS, spin dilution has led to greatly improved resolution, significant sensitivity enhancement and long-range 1H-1H correlations.[3, 4] Reif and coworkers have further obtained resolution rivalling solution NMR of larger proteins by back-exchanging with 10% H2O and 90% D2O.[5]

New methods for fast multidimensional NMR

Abstract: Considerable excitement has been aroused by recent new methods for speeding up multidimensional NMR experiments by radically modifying the normal time-domain sampling protocols. These new schemes include the filter diagonalization method, GFT-NMR, the single-scan two-dimensional technique, Hadamard spectroscopy, and a proposal based on projection-reconstruction of three-dimensional spectra. All these methods deliver appreciable improvements in the speed of data acquisition and show promise for speeding up multidimensional NMR of proteins. This perspective aims to describe these important new procedures in simple terms and to comment on their advantages and possible limitations.