Role of the amino-terminal residues of the interferon-induced protein kinase in its activation by double-stranded RNA and heparin.
We have previously reported that the amino-terminal residues 1-34 of the interferon-induced protein kinase (RNA-activated) (PKR) are necessary for its binding to and activation by double-stranded RNA (dsRNA) (Patel, R. C., and Sen, G. C. (1992) J. Biol. Chem. 267, 7671-7676). Here, we report that the amino-terminal 24 residues are indispensable for these properties of the enzyme. The replacement of these residues with 14 unrelated residues fully restored the protein's dsRNA binding activity, but only partially restored the enzyme activity. Mutation of residues 18 and 19 revealed their importance in determining the affinity of PKR for dsRNA and its ability to phosphorylate eukaryotic initiation factor 2 alpha. These mutations, however, did not affect PKR's autophosphorylation activity. Deletion mutants that failed to bind to and be activated by dsRNA could be fully activated by the alternative activator, heparin. Thus, activation of PKR by dsRNA and heparin is mediated through different mechanisms that require different domains of the protein.
Patel
R C
RC
Department of Molecular Biology, Cleveland Clinic Foundation, Ohio 44915-5285.
Stanton
P
P
Sen
G C
GC
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D003001:Cloning, Molecular; D004789:Enzyme Activation; D004790:Enzyme Induction; D015852:Eukaryotic Initiation Factor-2 / Q000378:metabolism; D006493:Heparin / Q000494:pharmacology*; D006801:Humans; D007372:Interferons / Q000494:pharmacology*; D051379:Mice; D008969:Molecular Sequence Data; D016296:Mutagenesis; D016297:Mutagenesis, Site-Directed; D009838:Oligodeoxyribonucleotides; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases / Q000096:biosynthesis / Q000302:isolation & purification / Q000378:metabolism*; D012330:RNA, Double-Stranded / Q000378:metabolism* / Q000494:pharmacology; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000302:isolation & purification / Q000378:metabolism; D017384:Sequence Deletion; D017386:Sequence Homology, Amino Acid; D019892:eIF-2 Kinase
D015852:Eukaryotic Initiation Factor-2; D009838:Oligodeoxyribonucleotides; D012330:RNA, Double-Stranded; D011994:Recombinant Proteins; D006493:Heparin; D007372:Interferons; D017346:Protein Serine-Threonine Kinases; D019892:eIF-2 Kinase