title: Characterization and structure of the Aquifex aeolicus protein DUF752: a bacterial tRNA-methyltransferase (MnmC2) functioning without the usually fused oxidase domain (MnmC1).
abstract: Post-transcriptional modifications of the wobble uridine (U34) of tRNAs play a critical role in reading NNA/G codons belonging to split codon boxes. In a subset of Escherichia coli tRNA, this wobble uridine is modified to 5-methylaminomethyluridine (mnm(5)U34) through sequential enzymatic reactions. Uridine 34 is first converted to 5-carboxymethylaminomethyluridine (cmnm(5)U34) by the MnmE-MnmG enzyme complex. The cmnm(5)U34 is further modified to mnm(5)U by the bifunctional MnmC protein. In the first reaction, the FAD-dependent oxidase domain (MnmC1) converts cmnm(5)U into 5-aminomethyluridine (nm(5)U34), and this reaction is immediately followed by the methylation of the free amino group into mnm(5)U34 by the S-adenosylmethionine-dependent domain (MnmC2). Aquifex aeolicus lacks a bifunctional MnmC protein fusion and instead encodes the Rossmann-fold protein DUF752, which is homologous to the methyltransferase MnmC2 domain of Escherichia coli MnmC (26% identity). Here, we determined the crystal structure of the A. aeolicus DUF752 protein at 2.5 Å resolution, which revealed that it catalyzes the S-adenosylmethionine-dependent methylation of nm(5)U in vitro, to form mnm(5)U34 in tRNA. We also showed that naturally occurring tRNA from A. aeolicus contains the 5-mnm group attached to the C5 atom of U34. Taken together, these results support the recent proposal of an alternative MnmC1-independent shortcut pathway for producing mnm(5)U34 in tRNAs.
authors.affiliation: RIKEN SPring-8 Center, Harima Institute, 1-1-1 Kouto, Sayo, Hyogo 679-5148, Japan.
authors.forename: Aya
authors.initials: A
authors.lastname: Kitamura
authors.forename: Madoka
authors.initials: M
authors.lastname: Nishimoto
authors.forename: Toru
authors.initials: T
authors.lastname: Sengoku
authors.forename: Rie
authors.initials: R
authors.lastname: Shibata
authors.forename: Gunilla
authors.initials: G
authors.lastname: Jäger
authors.forename: Glenn R
authors.initials: GR
authors.lastname: Björk
authors.forename: Henri
authors.initials: H
authors.lastname: Grosjean
authors.forename: Shigeyuki
authors.initials: S
authors.lastname: Yokoyama
authors.forename: Yoshitaka
authors.initials: Y
authors.lastname: Bessho
mesh_terms: D001419:Bacteria / Q000201:enzymology* / Q000235:genetics; D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D018360:Crystallography, X-Ray; D008745:Methylation; D010088:Oxidoreductases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D017434:Protein Structure, Tertiary; D012329:RNA, Bacterial / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D012343:RNA, Transfer / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D012436:S-Adenosylmethionine / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D012359:tRNA Methyltransferases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism
chemical_list: D001426:Bacterial Proteins; D012329:RNA, Bacterial; D012436:S-Adenosylmethionine; D012343:RNA, Transfer; D010088:Oxidoreductases; D012359:tRNA Methyltransferases