THE USE OF2H,13C,15N MULTIDIMENSIONAL NMR GTO STUDY THE STRUCTURE AND DYNAMICS OF PROTEINS

Gardner, Kevin H.; Kay, Lewis E.

doi:10.1146/annurev.biophys.27.1.357

Cited by 579 publications

(570 citation statements)

References 174 publications

(288 reference statements)

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“…In addition, as the number of protein residues increases, spectral overlap hinders analysis. Relatively novel techniques such as transverse relaxationoptimized spectroscopy (TROSY) experiments and per-deuteration of protein samples have advanced the protein size limit, whereas selective labeling has decreased the spectral overlap problem (12). Here, we use TCR and pMHC molecules whose sizes have been reduced to the point that NMR can be used quickly and economically to determine footprints of multiple TCR-pMHC complexes.…”

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confidence: 99%

Solution mapping of T cell receptor docking footprints on peptide-MHC

Varani¹,

Bankovich²,

Liu

et al. 2007

Proc. Natl. Acad. Sci. U.S.A.

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T cell receptor (TCR) recognition of peptide-MHC (pMHC) is central to the cellular immune response. A large database of TCR-pMHC structures is needed to reveal general structural principles, such as whether the repertoire of TCR/MHC docking modes is dictated by a ''recognition code'' between conserved elements of the TCR and MHC genes. Although Ϸ17 cocrystal structures of unique TCRpMHC complexes have been determined, cocrystallization of soluble TCR and pMHC remains a major technical obstacle in the field. Here we demonstrate a strategy, based on NMR chemical shift mapping, that permits rapid and reliable analysis of the solution footprint made by a TCR when binding onto the pMHC surface. We mapped the 2C TCR binding interaction with its allogeneic ligand H-2L d -QL9 and identified a group of NMR-shifted residues that delineated a clear surface of the MHC that we defined as the TCR footprint. We subsequently found that the docking footprint described by NMR shifts was highly accurate compared with a recently determined high-resolution crystal structure of the same complex. The same NMR footprint analysis was done on a highaffinity mutant of the TCR. The current work serves as a foundation to explore the molecular dynamics of pMHC complexes and to rapidly determine the footprints of many L d -specific TCRs.chemical shift mapping ͉ dynamics ͉ NMR ͉ cellular immunity ͉ protein-protein interaction T cells modulate the nature and extent of an immune response based primarily on specific T cell receptor (TCR) recognition of peptide in the context of a MHC molecule (1, 2). The TCR is a genetically recombined receptor, analogous to an antibody, that is composed of ␣ and ␤ chains. The binding site of the TCR comprises six loops called complementarity-determining regions (CDRs), with each chain contributing three loops, called CDR 1, 2, and 3. The CDR1 and CDR2 loop sequences are encoded by the variable (V) genes, but because there is no somatic mutation in TCR-V genes, these are referred to as ''germ line-derived.'' In contrast, the CDR3 loops, which are largely involved in antigenic peptide contact and thus specificity, are derived from recombination of V(D)J segments and vary in an almost unlimited fashion. MHC gene products present antigenic peptides to the TCR through a composite peptide-MHC (pMHC) surface composed of a highly variable element (i.e., peptide) in a groove surrounded by more conserved residues on the MHC helices (2).The interaction between TCR and pMHC has been studied through cocrystallization of Ϸ17 unique complexes (1, 3). Some general principles have emerged from the current database of complexes (4). Briefly, the TCR has an orientation over the pMHC surface that generally places the germline-encoded CDR1 and CDR2 loops in contact with the conserved helical residues of the MHC, whereas the highly variable, somatically recombined CDR3 loops primarily interact with the peptide. This docking orientation, also called the ''footprint,'' shows a high degree of variability (Ϯ60°) in different pMHC complex...

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mentioning

confidence: 99%

Solution mapping of T cell receptor docking footprints on peptide-MHC

Varani¹,

Bankovich²,

Liu

et al. 2007

Proc. Natl. Acad. Sci. U.S.A.

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“…In general, high-resolution NMR structures and assignments can be routinely obtained for proteins <25 kDa using standard 13 C and 15 N protein labeling techniques [32,33]. This molecular-weight upper limit may be extended by upwards of 900 kDa [34][35][36][37][38][39][40] by the application of deuterium labeling, specific methyl labeling and Transverse Relaxation-Optimized NMR Spectroscopy (TROSY)-based experiments [41][42][43].…”

Section: The Fast-nmr Methodsmentioning

confidence: 99%

The application of FAST-NMR for the identification of novel drug discovery targets

Powers

Mercier

Copeland

2008

Drug Discovery Today

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“…Instrumental and methodological advances in the past decades (Pervushin et al, 1997;Gardner and Kay, 1998;Tjandra and Bax, 1997;Orekhov et al, 2001) opened the possibilities for the investigation of proteins up to 100 kDa in the solution state (Tugarinov and Kay, 2003;Tugarinov et al, 2005), including the study of membrane proteins reconstituted in detergent micelles and bicelles (Sanders and Landis, 1995;Opella and Marassi, 2004). The most illustrative recent examples include the structure determination of a voltage-dependent anion channel from mitochondria, VDAC-1 (Hiller et al, 2008) and a GPCR-like seven-helical transmembrane receptor sensory rhodopsin II (Gautier et al, 2010).…”

Section: D Nuclear Magnetic Resonance (Nmr) Spectroscopymentioning

confidence: 99%

Asymmetric perturbations of signalling oligomers

Maksay

Tőke

2014

Progress in Biophysics and Molecular Biology

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This review focuses on rapid and reversible noncovalent interactions for symmetric oligomers of tetramers (voltage-gated cation channels, ionotropic glutamate receptor, CNG and CHN channels); pentameric ligand-gated and mechanosensitive channels; higher order oligomers (gap junction channel, chaperonins, proteasome, virus capsid); as well as primary and secondary transporters. In conclusion, asymmetric perturbations seem to play important functional roles in a broad range of communicating networks.

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THE USE OF²H,¹³C,¹⁵N MULTIDIMENSIONAL NMR GTO STUDY THE STRUCTURE AND DYNAMICS OF PROTEINS

Cited by 579 publications

References 174 publications

Solution mapping of T cell receptor docking footprints on peptide-MHC

Solution mapping of T cell receptor docking footprints on peptide-MHC

The application of FAST-NMR for the identification of novel drug discovery targets

Asymmetric perturbations of signalling oligomers

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