title: A single zinc ion is sufficient for an active Trypanosoma brucei tRNA editing deaminase.
abstract: Editing of adenosine (A) to inosine (I) at the first anticodon position in tRNA is catalyzed by adenosine deaminases acting on tRNA (ADATs). This essential reaction in bacteria and eukarya permits a single tRNA to decode multiple codons. Bacterial ADATa is a homodimer with two bound essential Zn(2+). The ADATa crystal structure revealed residues important for substrate binding and catalysis; however, such high resolution structural information is not available for eukaryotic tRNA deaminases. Despite significant sequence similarity among deaminases, we continue to uncover unexpected functional differences between Trypanosoma brucei ADAT2/3 (TbADAT2/3) and its bacterial counterpart. Previously, we demonstrated that TbADAT2/3 is unique in catalyzing two different deamination reactions. Here we show by kinetic analyses and inductively coupled plasma emission spectrometry that wild type TbADAT2/3 coordinates two Zn(2+) per heterodimer, but unlike any other tRNA deaminase, mutation of one of the key Zn(2+)-coordinating cysteines in TbADAT2 yields a functional enzyme with a single-bound zinc. These data suggest that, at least, TbADAT3 may play a role in catalysis via direct coordination of the catalytic Zn(2+). These observations raise the possibility of an unusual Zn(2+) coordination interface with important implications for the function and evolution of editing deaminases.
authors.affiliation: Department of Microbiology, The Ohio State Center for RNA Biology, Columbus, Ohio 43210, USA.
authors.forename: Jessica L
authors.initials: JL
authors.lastname: Spears
authors.forename: Mary Anne T
authors.initials: MA
authors.lastname: Rubio
authors.forename: Kirk W
authors.initials: KW
authors.lastname: Gaston
authors.forename: Ewa
authors.initials: E
authors.lastname: Wywial
authors.forename: Alexandros
authors.initials: A
authors.lastname: Strikoudis
authors.forename: Janusz M
authors.initials: JM
authors.lastname: Bujnicki
authors.forename: F Nina
authors.initials: FN
authors.lastname: Papavasiliou
authors.forename: Juan D
authors.initials: JD
authors.lastname: Alfonzo
mesh_terms: D000243:Adenosine Deaminase / Q000235:genetics / Q000378:metabolism*; D002413:Cations, Divalent / Q000378:metabolism; D015800:Protozoan Proteins / Q000235:genetics / Q000378:metabolism*; D017393:RNA Editing / Q000502:physiology*; D016053:RNA, Protozoan / Q000096:biosynthesis* / Q000235:genetics; D012343:RNA, Transfer / Q000096:biosynthesis* / Q000235:genetics; D016601:RNA-Binding Proteins; D014346:Trypanosoma brucei brucei / Q000201:enzymology* / Q000235:genetics; D015032:Zinc / Q000378:metabolism*
chemical_list: D002413:Cations, Divalent; D015800:Protozoan Proteins; D016053:RNA, Protozoan; D016601:RNA-Binding Proteins; D012343:RNA, Transfer; C567955:ADARB1 protein, human; D000243:Adenosine Deaminase; D015032:Zinc