Transverse Relaxation Optimized Spectroscopy TROSY

2 Transverse Relaxation Optimized Spectroscopy (TROSY). - TROSY and CRINEPT are new techniques for solution NMR studies of molecular and supramolecular structures. They allow the collection of high-resolution spectra of structures with molecular weights 100 kDa, significantly extending the range of macromolecular systems that can be studied by NMR in solution. TROSY has already been used to map protein-protein interfaces, to conduct structural studies on membrane proteins and to study nucleic acid conformations in multimolecular assemblies. A number of reviews have been published this year that cover the TROSY technique since its inception.  

The number of pulse sequences incorporating TROSY type magnetization transfer has steadily grown over the last year. The discovers of TROSY describe a N- H/ C- H-TROSY experiment for the simultaneous acquisition of the heteronuclear chemical shift correlations of backbone amide groups, side-chain N- H2 groups and aromatic groups.  

Recent developments in the direct observation of J couplings across hydrogen bonds in proteins and nucleic acids provide additional information for structure and function studies of these molecules by NMR spectroscopy. Yan et al proposed a modified J(N,N)-correlated TROSY experiment 

TROSY experiment for the measurement of three-bond scalar coupling constant between lH(i-l alpha) and 15N(i), which defines the dihedral angle. There are still relatively few examples of real applications where TROSY techniques have been essential to derive meaningful information. This is just a matter of time and this year has seen a few landmark illustrations. In a first 

The concept of chemical shift-coding monitors chemical shifts in multi-dimensional NMR experiments without additional polarization transfer elements and evolution periods. The chemical shifts are coded in the line-shape of the crosspeak through an apparent scalar coupling dependent upon the chemical shift. This concept has been applied to the 3D triple-resonance experiment HNCA adding the information of C(beta) or C chemical shifts. On average, the 

Most methods for determining residual dipolar couplings are based on the measurement of the displacement between cross-peak components in J-coupled spectra. However, for large macromolecules and macromolecular complexes, these methods are often unreliable since differential relaxation can significantly broaden one of the multiplet components and thereby make accurate determination of its position difficult. To overcome this problem, a J-evolved transverse relaxation optimized (JE-TROSY) method has been demonstrated for the determination of one-bond couplings that involves J-evolution of the sharpest crosspeak multiplet component selected in a TROSY experiment . Couplings are measured from the displacement of the TROSY component in the additional J-evolution dimension relative to a zero frequency origin. 

Relaxation in methyl groups is strongly influenced by cross-correlated interactions involving the methyl dipoles. One of the major interference effects results from intra-methyl H- C, H- H dipolar interactions and leads to significant 

The difference in the relaxation rates of ZQ and DQ coherences is the result of three principal mechanisms. These include the cross-correlation between the chemical shift anisotropies of the two participating nuclei, dipolar interactions with remote protons as well as interference effects due to the time-modulation of their isotropic chemical shifts as a consequence of slow mus-ms dynamics. The last effect when present, dominates the others resulting in large differences between the relaxation rates of ZQ and DQ coherences. Majumdar and Ghose have presented four TROSY-based experiments that measure this effect for several pairs of backbone nuclei. These experiments allow the detection of slow dynamic processes in the protein backbone including correlated motion over two and three bonds . A suite of TROSY-based NMR relaxation dispersion experiments that measure the decay of DG and ZQ coherences as a 

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New techniques for data analysis and improvements in instrumentation have now made it possible to carry out stmctural and conformational studies of biopolymers including proteins, polysaccharides, and nucleic acids. NMR, which may be done on noncrystalline materials in solution, provides a technique complementary to X-ray diffraction, which requires crystals for analysis. One-dimensional NMR, as described to this point, can offer structural data for smaller molecules. But proteins and other biopolymers with large numbers of protons will yield a very crowded spectrum with many overlapping lines. In multidimensional NMR (2-D, 3-D, 4-D), peaks are spread out through two or more axes to improve resolution. The techniques of correlation spectroscopy (COSY), nuclear Overhausser effect spectroscopy (NOESY), and transverse relaxation-optimized spectroscopy (TROSY) depend on the observation that nonequivalent protons interact with each other. By using multiple-pulse techniques, it is possible to perturb one nucleus and observe the effect on the spin states of other nuclei. The availability of powerful computers and Fourier transform (FT) calculations makes it possible to elucidate structures of proteins up to 40,000 daltons in molecular mass and there is future promise for studies on proteins over 100,000... 

NEW TECHNIQUES FOR PROTEIN NMR RESIDUAL DIPOLAR COUPLINGS AND TRANSVERSE RELAXATION OPTIMIZED SPECTROSCOPY (TROSY)... 

In liquid-state NMR, spin relaxation due to cross-correlation of two anisotropic spin interactions can provide useful information about molecular structure and dynamics. These effects are manifest as differential line widths or line intensities in the NMR spectra. Recently, new experiments were developed for the accurate measurement of numerous cross-correlated relaxation rates in scalar coupled multi-spin systems. The recently introduced concept of transverse relaxation optimized spectroscopy (TROSY) is also based on cross-correlated relaxation. Brutscher outlined the basic concepts and experimental techniques necessary for understanding and exploiting cross-correlated relaxation effects in macromolecules. In addition, he presented some examples showing the potential of cross-correlated relaxation for high-resolution NMR studies of proteins and nucleic acids. 

The most important recent advance of the solution NMR pulse sequence methodology is transverse relaxation optimized spectroscopy (TROSY) due to its significant sensitivity enhancement for large proteins by increasing the lifetimes of NMR signals.17 120 124-126... 

Structure determination. The introduction of transverse relaxation optimized spectroscopy (TROSY) by Wuthrich and co-workers [276] opened up a wealth of new opportunities for solution NMR on large protein systems, including detergent-solubilized membrane proteins. 

Pervushin K, Riek R, Wider G, Wiithrich K (1998) Transverse relaxation-optimized spectroscopy (TROSY) for NMR studies of aromatic spin systems in C-labeled proteins. J Am Chem Soc 120 6394-6400... 

A short overlook on transverse relaxation-optimized spectroscopy (TROSY) in combination with various isotope-labelling techniques has been published by Fernandez and Wider. The method improves, among others, the detection of scalar couplings across hydrogen bonds, providing crucial information on the determination of solution structures of large proteins and oligonucleotides. 

During the past two years, significant advances have been made in the development of NMR methods for studying biomolecular dynamics on the microsecond to millisecond timescale. Thuduppathy and HilP reviewed applications of NMR spin relaxation methods for measuring biological motions. Fernandez and Wider discussed transverse relaxation-optimized spectroscopy (TROSY) that promises to further enhance the determination of solution structures of large biomolecules. 

The relaxation interference between the H- N dipolar interaction and the nitrogen-15 CSA is the basis of the transverse relaxation optimized spectroscopy (TROSY), nowadays a standard tool in NMR of larger proteins. Zuiderweg and Rousaki reviewed the field of gradient-enhanced TROSY and described the experiments of this kind in terms of the cartesian product operators. Other midifications of TROSY have also been reported, but are judged to be beyond the scope of this review. 

Transverse relaxation-optimized spectroscopy (TROSY)-based NMR experiments for obtaining RDC s as well as other spectral parameters have been presented. A methodology that allows for the structural and dynamic... 

Transverse-relaxation optimized spectroscopy (TROSY) [5], as its name suggested, seeks to optimize (minimize) the relaxation of transverse signals. Transverse signals are the observable (directly or indirectly) signals in modem NMR,... 

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