CCR5 RNA Pseudoknots: Residue and Site-Specific Labeling correlate Internal Motions with microRNA Binding.
Chemistry
; 24(21): 5462-5468, 2018 Apr 11.
Article
em En
| MEDLINE
| ID: mdl-29412477
ABSTRACT
Conformational dynamics of RNA molecules play a critical role in governing their biological functions. Measurements of RNA dynamic behavior sheds important light on sites that interact with their binding partners or cellular stimulators. However, such measurements using solution-state NMR are difficult for large RNA molecules (>70â
nt; nt=nucleotides) owing to severe spectral overlap, homonuclear 13 C scalar couplings, and line broadening. Herein, a strategic combination of solid-phase synthesis, site-specific isotopic labeled phosphoramidites, and enzymatic ligation is introduced. This approach allowed the position-specific insertion of isotopic probes into a 96â
nt CCR5 RNA fragment. Accurate measurements of functional dynamics using the Carr-Purcell-Meiboom-Gill (CPMG) relaxation dispersion (RD) experiments enabled extraction of the exchange rates and populations of this RNA. NMR chemical shift perturbation analysis of the RNA/microRNA-1224 complex indicated that A90-C1' of the pseudoknot exhibits similar changes in chemical shift observed in the excited state. This work demonstrates the general applicability of a NMR-labeling strategy to probe functional RNA structural dynamics.
Palavras-chave
Texto completo:
1
Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Modelos Moleculares
/
Receptores CCR5
/
MicroRNAs
Idioma:
En
Revista:
Chemistry
Assunto da revista:
QUIMICA
Ano de publicação:
2018
Tipo de documento:
Article
País de afiliação:
Estados Unidos