title: Synthesis and evaluation of an RNA editing substrate bearing 2'-deoxy-2'-mercaptoadenosine.
abstract: The RNA-editing adenosine deaminases (ADARs) catalyze deamination of adenosine to inosine in double stranded structure found in various RNA substrates, including mRNAs. Here we describe the synthesis of a phosphoramidite of 2'-deoxy-2'-mercaptoadenosine and its incorporation into an ADAR substrate. Surprisingly, no deamination product was observed with this substrate indicating replacing the 2'-OH with a 2'-SH at the editing site is highly inhibitory. Modeling of nucleotide binding into the active site suggests the side chain of T375 of human ADAR2 to be in proximity of the 2'-substituent. Mutation of this residue to cysteine caused a greater that 100-fold reduction in deamination rate with the 2'-OH substrate.
authors.affiliation: Department of Chemistry, University of California Davis, California 95616, USA.
authors.forename: Prasanna
authors.initials: P
authors.lastname: Jayalath
authors.forename: Subhash
authors.initials: S
authors.lastname: Pokharel
authors.forename: Eduardo
authors.initials: E
authors.lastname: Véliz
authors.forename: Peter A
authors.initials: PA
authors.lastname: Beal
mesh_terms: D000241:Adenosine / Q000031:analogs & derivatives* / Q000138:chemical synthesis / Q000737:chemistry / Q000378:metabolism; D000243:Adenosine Deaminase / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D001665:Binding Sites / Q000235:genetics; D003545:Cysteine / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D006801:Humans; D009843:Oligoribonucleotides / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D017393:RNA Editing; D016601:RNA-Binding Proteins; D013379:Substrate Specificity
chemical_list: D009843:Oligoribonucleotides; D016601:RNA-Binding Proteins; C567955:ADARB1 protein, human; D000243:Adenosine Deaminase; D000241:Adenosine; D003545:Cysteine