Protein kinase PKR mutants resistant to the poxvirus pseudosubstrate K3L protein.
Proc Natl Acad Sci U S A
; 105(44): 16894-9, 2008 Nov 04.
Article
en En
| MEDLINE
| ID: mdl-18971339
ABSTRACT
As part of the mammalian cell innate immune response, the double-stranded RNA activated protein kinase PKR phosphorylates the translation initiation factor eIF2alpha to inhibit protein synthesis and thus block viral replication. Poxviruses including vaccinia and smallpox viruses express PKR inhibitors such as the vaccinia virus K3L protein that resembles the N-terminal substrate-targeting domain of eIF2alpha. Whereas high-level expression of human PKR was toxic in yeast, this growth inhibition was suppressed by coexpression of the K3L protein. We used this yeast assay to screen for PKR mutants that are resistant to K3L inhibition, and we identified 12 mutations mapping to the C-terminal lobe of the PKR kinase domain. The PKR mutations specifically conferred resistance to the K3L protein both in yeast and in vitro. Consistently, the PKR-D486V mutation led to nearly a 15-fold decrease in K3L binding affinity yet did not impair eIF2alpha phosphorylation. Our results support the identification of the eIF2alpha-binding site on an extensive face of the C-terminal lobe of the kinase domain, and they indicate that subtle changes to the PKR kinase domain can drastically impact pseudosubstrate inhibition while leaving substrate phosphorylation intact. We propose that these paradoxical effects of the PKR mutations on pseudosubstrate vs. substrate interactions reflect differences between the rigid K3L protein and the plastic nature of eIF2alpha around the Ser-51 phosphorylation site.
Texto completo:
1
Colección:
01-internacional
Base de datos:
MEDLINE
Contexto en salud:
4_TD
Problema de salud:
4_smallpox
Asunto principal:
Proteínas Virales
/
EIF-2 Quinasa
Tipo de estudio:
Prognostic_studies
Idioma:
En
Revista:
Proc Natl Acad Sci U S A
Año:
2008
Tipo del documento:
Article
País de afiliación:
Estados Unidos