By providing a quality measure of the fit of the derived structure, it is analogous to the R-factor used for assessing structures derived using crystallography. The comparison of simulated and measured NOESY

spectra allows an estimate of the magnitude and direction of changes to be made to the molecule that might improve the agreement between the spectra. In order to achieve the full benefit of back-calculation, it is necessary to make it an integral part of the strategy for protein structure determination. This would involve a Dolutegravir datasheet readjustment of the distance restrains used in the structure calculation steps after analyzing the calculated NOESY spectrum. A new structure would be calculated and the process repeated until simulated and measured spectra match. For structure determination on the basis of distance constraints such as distance geometry and constrained find more molecular dynamics, among others, specialized softwares like NMRchitect can be used. The validity of the NMR method was established

conclusively by determining the three dimensional structure of the protein “tendamist” independently using NMR and normal X-ray diffraction analysis (Billiter et al., 1989). At present the use of 1H, 13C and 15N labeled proteins, three- and four-dimensional heteronuclear NMR spectroscopy together with TROSY offer a way to improve spectral resolution and circumvent problems due to larger line widths that are associated with increasing molecular weight. With these methodologies the determination of a high resolution NMR structure of proteins in the range of 100 kDa has been made possible (for review see Tugarinov et al., 2004). As discussed before NMR spectroscopy is a useful tool for studying one of the most important issues in biology, the

interaction of ligands with macromolecules. When part of the macromolecule is in close proximity to a bound ligand, a NOE can be observed in the ligand if the protons in Y-27632 2HCl the macromolecule are irradiated (James and Cohn, 1974). Concomitant with the developments in two-dimensional NMR and the use of NMR to determine the structure of peptides and proteins in solution, interest in transfer NOE (TRNOE) emerged (Cambell and Sykes, 1993). TRNOE is the extension of two dimensional NOE to exchanging systems such as ligand–protein complexes. TRNOE measurements give information on the conformation of the bound ligand. This methodology has been used to study the conformations of nucleotides bound to peptides and proteins (Leanz and Hammes, 1986 and Koide et al., 1989), binding of peptides to phospholipid bilayers (Milon et al., 1990), the codon to anticodon interaction (Clore et al., 1984), binding of peptides to enzymes (Meyer et al., 1988), binding of hormones to proteins (Live et al., 1987), drug discovery (Lucas et al., 2003) and binding of ligands to enzymes (Kuntothom et al., 2010). This methodology is used to characterize the binding of a ligand to a macromolecule at atomic resolution.