Pinpointing RNA-Protein Cross-Links with Site-Specific Stable Isotope-Labeled Oligonucleotides.
J Am Chem Soc
; 137(49): 15378-81, 2015 Dec 16.
Article
en En
| MEDLINE
| ID: mdl-26583201
ABSTRACT
High affinity RNA-protein interactions are critical to cellular function, but directly identifying the determinants of binding within these complexes is often difficult. Here, we introduce a stable isotope mass labeling technique to assign specific interacting nucleotides in an oligonucleotide-protein complex by photo-cross-linking. The method relies on generating site-specific oxygen-18-labeled phosphodiester linkages in oligonucleotides, such that covalent peptide-oligonucleotide cross-link sites arising from ultraviolet irradiation can be assigned to specific sequence positions in both RNA and protein simultaneously by mass spectrometry. Using Lin28A and a let-7 pre-element RNA, we demonstrate that mass labeling permits unambiguous identification of the cross-linked sequence positions in the RNA-protein complex.
Texto completo:
1
Colección:
01-internacional
Banco de datos:
MEDLINE
Asunto principal:
Oligonucleótidos
/
ARN
/
Proteínas
/
Reactivos de Enlaces Cruzados
Idioma:
En
Revista:
J Am Chem Soc
Año:
2015
Tipo del documento:
Article
País de afiliación:
Estados Unidos