title: Structural basis of AdoMet-dependent aminocarboxypropyl transfer reaction catalyzed by tRNA-wybutosine synthesizing enzyme, TYW2.
abstract: S-adenosylmethionine (AdoMet) is a methyl donor used by a wide variety of methyltransferases, and it is also used as the source of an alpha-amino-alpha-carboxypropyl ("acp") group by several enzymes. tRNA-yW synthesizing enzyme-2 (TYW2) is involved in the biogenesis of a hypermodified nucleotide, wybutosine (yW), and it catalyzes the transfer of the "acp" group from AdoMet to the C7 position of the imG-14 base, a yW precursor. This modified nucleoside yW is exclusively located at position 37 of eukaryotic tRNA(Phe), and it ensures the anticodon-codon pairing on the ribosomal decoding site. Although this "acp" group has a significant role in preventing decoding frame shifts, the mechanism of the "acp" group transfer by TYW2 remains unresolved. Here we report the crystal structures and functional analyses of two archaeal homologs of TYW2 from Pyrococcus horikoshii and Methanococcus jannaschii. The in vitro mass spectrometric and radioisotope-labeling analyses confirmed that these archaeal TYW2 homologues have the same activity as yeast TYW2. The crystal structures verified that the archaeal TYW2 contains a canonical class-I methyltransferase (MTase) fold. However, their AdoMet-bound structures revealed distinctive AdoMet-binding modes, in which the "acp" group, instead of the methyl group, of AdoMet is directed to the substrate binding pocket. Our findings, which were confirmed by extensive mutagenesis studies, explain why TYW2 transfers the "acp" group, and not the methyl group, from AdoMet to the nucleobase.
authors.affiliation: Department of Basic Medical Sciences, Institute of Medical Science, The University of Tokyo, 4-6-1 Shirokanedai, Minato-ku, Tokyo 108-8639, Japan.
authors.forename: Masataka
authors.initials: M
authors.lastname: Umitsu
authors.forename: Hiroshi
authors.initials: H
authors.lastname: Nishimasu
authors.forename: Akiko
authors.initials: A
authors.lastname: Noma
authors.forename: Tsutomu
authors.initials: T
authors.lastname: Suzuki
authors.forename: Ryuichiro
authors.initials: R
authors.lastname: Ishitani
authors.forename: Osamu
authors.initials: O
authors.lastname: Nureki
mesh_terms: D020134:Catalytic Domain; D018360:Crystallography, X-Ray; D017017:Methanococcus / Q000201:enzymology / Q000235:genetics; D008958:Models, Molecular; D009705:Nucleosides / Q000096:biosynthesis*; D043902:Pyrococcus horikoshii / Q000201:enzymology / Q000235:genetics; D012323:RNA Processing, Post-Transcriptional; D019642:RNA, Archaeal / Q000737:chemistry / Q000378:metabolism; D012360:RNA, Transfer, Phe / Q000737:chemistry / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D012436:S-Adenosylmethionine / Q000378:metabolism*; D055672:Static Electricity; D012359:tRNA Methyltransferases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*
chemical_list: D009705:Nucleosides; D019642:RNA, Archaeal; D012360:RNA, Transfer, Phe; D011994:Recombinant Proteins; C467591:wybutosine; D012436:S-Adenosylmethionine; D012359:tRNA Methyltransferases