DOTA-amide lanthanide tag for reliable generation of pseudocontact shifts in protein NMR spectra.
Bioconjug Chem
; 22(10): 2118-25, 2011 Oct 19.
Article
en En
| MEDLINE
| ID: mdl-21877751
ABSTRACT
Structural studies of proteins and protein-ligand complexes by nuclear magnetic resonance (NMR) spectroscopy can be greatly enhanced by site-specific attachment of lanthanide ions to create paramagnetic centers. In particular, pseudocontact shifts (PCS) generated by paramagnetic lanthanides contain important and unique long-range structure information. Here, we present a high-affinity lanthanide binding tag that can be attached to single cysteine residues of proteins. The new tag has many advantageous features that are not available in this combination from previously published tags (i) it binds lanthanide ions very tightly, minimizing the generation of nonspecific effects, (ii) it produces PCSs with high reliability as its bulkiness prevents complete motional averaging of PCSs, (iii) it can be attached to single cysteine residues, alleviating the need of detailed prior knowledge of the 3D structure of the target protein, and (iv) it does not display conformational exchange phenomena that would increase the number of signals in the NMR spectrum. The performance of the tag is demonstrated with the N-terminal domain of the E. coli arginine repressor and the A28C mutant of human ubiquitin.
Texto completo:
1
Bases de datos:
MEDLINE
Asunto principal:
Proteínas
/
Resonancia Magnética Nuclear Biomolecular
/
Elementos de la Serie de los Lantanoides
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Compuestos Heterocíclicos con 1 Anillo
Tipo de estudio:
Prognostic_studies
Límite:
Humans
Idioma:
En
Revista:
Bioconjug Chem
Asunto de la revista:
BIOQUIMICA
Año:
2011
Tipo del documento:
Article
País de afiliación:
Australia