Mechanism of interferon action: identification of a RNA binding domain within the N-terminal region of the human RNA-dependent P1/eIF-2 alpha protein kinase.
A molecular cDNA clone of the human RNA-dependent P1/eIF-2 alpha protein kinase was expressed in Escherichia coli. Mutant P1 proteins were examined for RNA binding activity by Northwestern blot analysis using the reovirus s1 mRNA, an activator of the kinase; the adenovirus VAI RNA, an inhibitor of kinase activation; or human immunodeficiency virus (HIV) TAR RNA as probe. Analysis of TrpE-P1 deletion mutant fusion proteins revealed that the 11-kDa N-terminal region of the P1 protein bound reovirus s1 mRNA, adenovirus VAI RNA, and HIV TAR RNA. Neither s1 RNA, VAI RNA, nor TAR RNA was bound by truncated P1 proteins which lacked the N-terminal 98 amino acids. Computer analysis revealed that the human protein P1 sequence corresponding to amino acid residues within the N-terminal RNA binding domain displays high homology (greater than 54% identity; 61 to 94% similarity) with two animal virus proteins which possess RNA binding activity (vaccinia virus E3L; rotavirus VP2) and two proteins of unknown function (murine TIK; rotavirus NS34), but which are likely RNA binding proteins.
McCormack
S J
SJ
Department of Biological Sciences, University of California, Santa Barbara 93106.
Thomis
D C
DC
Samuel
C E
CE
D000595:Amino Acid Sequence; D001483:Base Sequence; D001665:Binding Sites; D015153:Blotting, Western; D002872:Chromosome Deletion; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics; D015852:Eukaryotic Initiation Factor-2 / Q000378:metabolism; D006801:Humans; D007372:Interferons / Q000494:pharmacology*; D008969:Molecular Sequence Data; D011487:Protein Conformation; D011494:Protein Kinases / Q000235:genetics / Q000378:metabolism*; D012367:RNA, Viral / Q000378:metabolism*; D016601:RNA-Binding Proteins / Q000235:genetics*; D012689:Sequence Homology, Nucleic Acid; D019892:eIF-2 Kinase
D015852:Eukaryotic Initiation Factor-2; D012367:RNA, Viral; D016601:RNA-Binding Proteins; D007372:Interferons; D011494:Protein Kinases; D019892:eIF-2 Kinase