Biophysical characterizations of the recognition of the AAUAAA polyadenylation signal.
RNA
; 25(12): 1673-1680, 2019 12.
Article
en En
| MEDLINE
| ID: mdl-31462423
ABSTRACT
Most eukaryotic messenger RNA precursors must undergo 3'-end cleavage and polyadenylation for maturation. We and others recently reported the structure of the AAUAAA polyadenylation signal (PAS) in complex with the protein factors CPSF-30, WDR33, and CPSF-160, revealing the molecular mechanism for this recognition. Here we have characterized in detail the interactions between the PAS RNA and the protein factors using fluorescence polarization experiments. Our studies show that AAUAAA is recognized with â¼3 nM affinity by the CPSF-160-WDR33-CPSF-30 ternary complex. Variations in the RNA sequence can greatly reduce the affinity. Similarly, mutations of CPSF-30 residues that have van der Waals interactions with the bases of AAUAAA also lead to substantial reductions in affinity. Finally, our studies confirm that both CPSF-30 and WDR33 are required for high-affinity binding of the PAS RNA, while these two proteins alone and their binary complexes with CPSF-160 have much lower affinity for the RNA.
Palabras clave
Texto completo:
1
Colección:
01-internacional
Banco de datos:
MEDLINE
Asunto principal:
Poli A
/
ARN Mensajero
/
Proteínas Nucleares
/
Precursores del ARN
/
Poliadenilación
/
Factores de Escisión y Poliadenilación de ARNm
Límite:
Humans
Idioma:
En
Revista:
RNA
Asunto de la revista:
BIOLOGIA MOLECULAR
Año:
2019
Tipo del documento:
Article
País de afiliación:
Estados Unidos