Proteins spectral assignment

This example demonstrates how cell-free protein expression can be used to rapidly optimize the reaction conditions as well as to shorten the time for spectral assignments by producing individually labeled proteins. 

The spectral assignment process, complex as it is, is only a prelude to the use of NMR data to assist in the final determination of the three-dimensional structure of the protein. Here, the most important experiment is NOESY, often in conjunction with HMQC or other methods for spreading out the 2D NOESY peaks in three or four dimensions. For larger proteins 4D experiments are essential to spread the NOESY peaks according to both 13C and 15N chemical shifts. We... 

Once a suitable membrane protein sample is prepared, spectral assignments are made. Assignment is another rate-limiting step in NMR structure determination, and it has greatly benefited... 

Petkova and Tycko have described investigations of the effects of rotational resonance (R ) on solid state C NMR spectra of uniformly C-labelled samples obtained under MAS." The utility of R measurements as structural probes of peptides and proteins with multiple uniformly labelled residues has been demonstrated. Results for uniformly C-labelled L-alanine and L-valine in polycrystalline form and for amyloid fibrils formed by the 15-residue peptide with uniform labeling of a four-residue segment have been reported. The MAS NMR spectra revealed a novel J-decoupling effect at R conditions that may be useful in spectral assignments for systems with sharp C MAS NMR lines. Pronounced dependences of the apparent isotropic NMR chemical shifts on MAS frequency near R conditions were also observed. The feasibility of quantitative distance determinations in L-valine, and qualitative determinations of inter-residue contacts in fibrils has been demonstrated. In addition, it has... 

Infrared spectroscopy has been used in a large number of studies of proteins in a range of environments [22] and the spectral assignments are based on the fact that the secondary structures of these globular proteins have been very well... 

Interestingly, the behaviour of the Overhauser factor can be exploited to provide information about the spectral assignment and molecular dynamics of complex biological species. Thus, the intensity variations displayed by the decoupled and coupled spectra of whole cells isotopi-cally labelled is very illustrative (L 40). Whereas the resonances of the protein backbone amides are clearly observed as a broad signal at about 255 ppm in a proton non-decoupled spectrum measured at 9.22 MHz, the envelope of these resonances is nulled by proton broad-band decoupling. This result indicates either a reorientation rate of the cellular... 

Raman spectroscopy is a suitable and direct technique that provides information on secondary and tertiary protein structures [23-26]. It is a powerful tool for investigation of protein structure in solid and liquid food systems in general [14, 27, 28], and for determination of the protein structure in muscle foods (meat or fish) in particular [15-18]. The spectral assignments of protein Raman bands are usually based on model compoxmds such as amino acids or short peptides. 

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