Pure shift amide detection in conventional and TROSY-type experiments of C,N-labeled proteins.

Large coupling networks in uniformly C,N-labeled biomolecules induce broad multiplets that even in flexible proteins are frequently not recognized as such. The reason is that given multiplets typically consist of a large number of individual resonances that result in a single broad line, in which individual components are no longer resolved. We here introduce a real-time pure shift acquisition scheme for the detection of amide protons which is based on C-BIRD.

SORDOR pulses: expansion of the Böhlen-Bodenhausen scheme for low-power broadband magnetic resonance.

A novel type of efficient broadband pulse, called second-order phase dispersion by optimised rotation (SORDOR), has recently been introduced. In contrast to adiabatic excitation, SORDOR-90 pulses provide effective transverse 90 rotations throughout their bandwidth, with a quadratic offset dependence of the phase in the plane. Together with phase-matched SORDOR-180 pulses, this enables the Böhlen-Bodenhausen broadband refocusing approach for linearly frequency-swept pulses to be extended to any type of 90/180 pulse-delay sequence.

Concurrent J-evolving refocusing pulses.

Conventional refocusing pulses are optimised for a single spin without considering any type of coupling. However, despite the fact that most couplings will result in undesired distortions, refocusing in delay-pulse-delay-type sequences with desired heteronuclear coherence transfer might be enhanced considerably by including coupling evolution into pulse design. We provide a proof of principle study for a Hydrogen-Carbon refocusing pulse sandwich with inherent J-evolution following the previously reported ICEBERG-principle with improved performance in terms of refocusing performance and/or overall effective coherence transfer time.

Power of Pure Shift HαCα Correlations: A Way to Characterize Biomolecules under Physiological Conditions.

Intrinsically disordered proteins (IDPs) constitute an important class of biomolecules with high flexibility. Atomic-resolution studies for these molecules are essentially limited to NMR spectroscopy, which should be performed under physiological pH and temperature to populate relevant conformational ensembles. In this context, however, fundamental problems arise with established triple resonance NMR experiments: high solvent accessibility of IDPs promotes water exchange, which disfavors classical amide H-detection, while C-detection suffers from significantly reduced sensitivity.

Real-time pure shift measurements for uniformly isotope-labeled molecules using X-selective BIRD homonuclear decoupling.

We introduce a novel selective inversion element for chunked homonuclear decoupling that combines isotope selection via BIRD-filtering with band-selective inversion on the X-heteronucleus and allows efficient real-time decoupling of homonuclear and heteronuclear couplings. It is especially suitable for uniformly isotope-labeled compounds. We discuss in detail the inversion element based on band-selective refocusing on the X-nuclei (BASEREX), highlighting in particular the role of appropriate band-selective shaped refocusing pulses and the application of broadband X-pulses for an effective BIRD element during homodecoupling.

Boosting the NMR Assignment of Carbohydrates with Clean In-Phase Correlation Experiments.

Novel CLIP-COSY based homo- and heteronuclear correlation experiments are reported for the rapid, semi-automated NMR assignment of small to medium-sized molecules. The homonuclear CLIP-COSY and corresponding relayed experiments employ the perfect-echo based mixing sequence for in-phase coherence transfer between directly and/or indirectly coupled proton spins. The combined analysis of the resulting CLIP-COSY and relayed spectra made it possible to easily track down, layer by layer, the proton-proton connectivity network.

Observing the overall rocking motion of a protein in a crystal.

The large majority of three-dimensional structures of biological macromolecules have been determined by X-ray diffraction of crystalline samples. High-resolution structure determination crucially depends on the homogeneity of the protein crystal. Overall 'rocking' motion of molecules in the crystal is expected to influence diffraction quality, and such motion may therefore affect the process of solving crystal structures.

Probing transient conformational states of proteins by solid-state R(1ρ) relaxation-dispersion NMR spectroscopy.

The function of proteins depends on their ability to sample a variety of states differing in structure and free energy. Deciphering how the various thermally accessible conformations are connected, and understanding their structures and relative energies is crucial in rationalizing protein function. Many biomolecular reactions take place within microseconds to milliseconds, and this timescale is therefore of central functional importance.

Amplitudes and time scales of picosecond-to-microsecond motion in proteins studied by solid-state NMR: a critical evaluation of experimental approaches and application to crystalline ubiquitin.

Solid-state NMR provides insight into protein motion over time scales ranging from picoseconds to seconds. While in solution state the methodology to measure protein dynamics is well established, there is currently no such consensus protocol for measuring dynamics in solids. In this article, we perform a detailed investigation of measurement protocols for fast motions, i.