title: Evidence for auto-inhibition by the N terminus of hADAR2 and activation by dsRNA binding.
abstract: Adenosine deaminases that act on RNA (ADARs) catalyze adenosine to inosine conversion in RNA that is largely double stranded. Human ADAR2 (hADAR2) contains two double-stranded RNA binding motifs (dsRBMs), separated by a 90-amino acid linker, and these are followed by the C-terminal catalytic domain. We assayed enzymatic activity of N-terminal deletion constructs of hADAR2 to determine the role of the dsRBMs and the intervening linker peptide. We found that a truncated protein consisting of one dsRBM and the deaminase domain was capable of deaminating a short 15-bp substrate. In contrast, full-length hADAR2 was inactive on this short substrate. In addition, we observed that the N terminus, which was deleted from the truncated protein, inhibits editing activity when added in trans. We propose that the N-terminal domain of hADAR2 contains sequences that cause auto-inhibition of the enzyme. Our results suggest activation requires binding to an RNA substrate long enough to accommodate interactions with both dsRBMs.
authors.affiliation: Howard Hughes Medical Institute and Department of Biochemistry, University of Utah, Salt Lake City, UT 84132, USA.
authors.forename: Mark R
authors.initials: MR
authors.lastname: Macbeth
authors.forename: Arunth T
authors.initials: AT
authors.lastname: Lingam
authors.forename: Brenda L
authors.initials: BL
authors.lastname: Bass
mesh_terms: D000243:Adenosine Deaminase / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D058892:Adenosine Deaminase Inhibitors; D000595:Amino Acid Sequence; D001483:Base Sequence; D020134:Catalytic Domain; D004789:Enzyme Activation; D006801:Humans; D066298:In Vitro Techniques; D008954:Models, Biological; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D017434:Protein Structure, Tertiary; D012313:RNA / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D017393:RNA Editing; D016601:RNA-Binding Proteins; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D017384:Sequence Deletion
chemical_list: D058892:Adenosine Deaminase Inhibitors; D016601:RNA-Binding Proteins; D011994:Recombinant Proteins; D012313:RNA; C567955:ADARB1 protein, human; D000243:Adenosine Deaminase