Dynamic flexibility of double-stranded RNA activated PKR in solution.
PKR, an interferon-induced double-stranded RNA activated serine-threonine kinase, is a component of signal transduction pathways mediating cell growth control and responses to stress and viral infection. Analysis of separate PKR functional domains by NMR and X-ray crystallography has revealed details of PKR RNA binding domains and kinase domain, respectively. Here, we report the structural characteristics, calculated from biochemical and neutron scattering data, of a native PKR fraction with a high level of autophosphorylation and constitutive kinase activity. The experiments reveal association of the protein monomer into dimers and tetramers, in the absence of double-stranded RNA or other activators. Low-resolution structures of the association states were obtained from the large angle neutron scattering data and reveal the relative orientation of all protein domains in the activated kinase dimer. Low-resolution structures were also obtained for a PKR tetramer-monoclonal antibody complex. Taken together, this information leads to a new model for the structure of the functioning unit of the enzyme, highlights the flexibility of PKR and sheds light on the mechanism of PKR activation. The results of this study emphasize the usefulness of low-resolution structural studies in solution on large flexible multiple domain proteins.
Gabel
Frank
F
Structural and Computational Biology Unit, European Molecular Biology Laboratory, 69117 Heidelberg, Germany.
Wang
Die
D
Madern
Dominique
D
Sadler
Anthony
A
Dayie
Kwaku
K
Daryoush
Maryam Zamanian
MZ
Schwahn
Dietmar
D
Zaccai
Giuseppe
G
Lee
Xavier
X
Williams
Bryan R G
BR
D001704:Biopolymers; D018360:Crystallography, X-Ray; D004789:Enzyme Activation; D006801:Humans; D007142:Immunoglobulin gamma-Chains / Q000737:chemistry; D008958:Models, Molecular; D019906:Nuclear Magnetic Resonance, Biomolecular; D010766:Phosphorylation; D011485:Protein Binding; D011487:Protein Conformation; D012330:RNA, Double-Stranded / Q000737:chemistry*; D012996:Solutions; D019892:eIF-2 Kinase / Q000737:chemistry*
D001704:Biopolymers; D007142:Immunoglobulin gamma-Chains; D012330:RNA, Double-Stranded; D012996:Solutions; D019892:eIF-2 Kinase