title: The non-canonical hydroxylase structure of YfcM reveals a metal ion-coordination motif required for EF-P hydroxylation.
abstract: EF-P is a bacterial tRNA-mimic protein, which accelerates the ribosome-catalyzed polymerization of poly-prolines. In Escherichia coli, EF-P is post-translationally modified on a conserved lysine residue. The post-translational modification is performed in a two-step reaction involving the addition of a β-lysine moiety and the subsequent hydroxylation, catalyzed by PoxA and YfcM, respectively. The β-lysine moiety was previously shown to enhance the rate of poly-proline synthesis, but the role of the hydroxylation is poorly understood. We solved the crystal structure of YfcM and performed functional analyses to determine the hydroxylation mechanism. In addition, YfcM appears to be structurally distinct from any other hydroxylase structures reported so far. The structure of YfcM is similar to that of the ribonuclease YbeY, even though they do not share sequence homology. Furthermore, YfcM has a metal ion-coordinating motif, similar to YbeY. The metal ion-coordinating motif of YfcM resembles a 2-His-1-carboxylate motif, which coordinates an Fe(II) ion and forms the catalytic site of non-heme iron enzymes. Our findings showed that the metal ion-coordinating motif of YfcM plays an essential role in the hydroxylation of the β-lysylated lysine residue of EF-P. Taken together, our results suggested the potential catalytic mechanism of hydroxylation by YfcM.
authors.affiliation: Department of Biological Sciences, Graduate School of Science, The University of Tokyo, 2-11-16 Yayoi, Bunkyo-ku, Tokyo 113-0032, Japan Global Research Cluster, RIKEN, 2-1, Hirosawa, Wako, Saitama, 351-0198, Japan.
authors.forename: Kan
authors.initials: K
authors.lastname: Kobayashi
authors.affiliation: Department of Microbiology, Ohio State University, Columbus, OH 43210, USA.
authors.forename: Assaf
authors.initials: A
authors.lastname: Katz
authors.affiliation: Molecular, Cell, and Developmental Biology Program, Ohio State University, Columbus, OH 43210, USA.
authors.forename: Andrei
authors.initials: A
authors.lastname: Rajkovic
authors.affiliation: Department of Biological Sciences, Graduate School of Science, The University of Tokyo, 2-11-16 Yayoi, Bunkyo-ku, Tokyo 113-0032, Japan Global Research Cluster, RIKEN, 2-1, Hirosawa, Wako, Saitama, 351-0198, Japan.
authors.forename: Ryohei
authors.initials: R
authors.lastname: Ishii
authors.affiliation: Department of Biochemistry, Ohio State University, Columbus, OH 43210, USA.
authors.forename: Owen E
authors.initials: OE
authors.lastname: Branson
authors.affiliation: Comprehensive Cancer Center, Ohio State University, Columbus, OH 43210, USA Department of Molecular Virology, Immunology and Medical Genetics, Ohio State University, Columbus, OH 43210, USA.
authors.forename: Michael A
authors.initials: MA
authors.lastname: Freitas
authors.affiliation: Department of Biological Sciences, Graduate School of Science, The University of Tokyo, 2-11-16 Yayoi, Bunkyo-ku, Tokyo 113-0032, Japan Global Research Cluster, RIKEN, 2-1, Hirosawa, Wako, Saitama, 351-0198, Japan.
authors.forename: Ryuichiro
authors.initials: R
authors.lastname: Ishitani
authors.affiliation: Department of Microbiology, Ohio State University, Columbus, OH 43210, USA Ohio State Biochemistry Program, Center for RNA Biology, Ohio State University, Columbus, OH 43210, USA.
authors.forename: Michael
authors.initials: M
authors.lastname: Ibba
authors.affiliation: Department of Biological Sciences, Graduate School of Science, The University of Tokyo, 2-11-16 Yayoi, Bunkyo-ku, Tokyo 113-0032, Japan Global Research Cluster, RIKEN, 2-1, Hirosawa, Wako, Saitama, 351-0198, Japan nureki@bs.s.u-tokyo.ac.jp.
authors.forename: Osamu
authors.initials: O
authors.lastname: Nureki
mesh_terms: D020816:Amino Acid Motifs; D029968:Escherichia coli Proteins / Q000737:chemistry* / Q000378:metabolism; D006900:Hydroxylation; D008667:Metalloproteins / Q000737:chemistry; D008670:Metals / Q000737:chemistry*; D006899:Mixed Function Oxygenases / Q000737:chemistry* / Q000378:metabolism; D008958:Models, Molecular; D010445:Peptide Elongation Factors / Q000378:metabolism*; D011499:Protein Processing, Post-Translational
chemical_list: D029968:Escherichia coli Proteins; D008667:Metalloproteins; D008670:Metals; D010445:Peptide Elongation Factors; C510617:YbeY protein, E coli; C020746:factor EF-P; D006899:Mixed Function Oxygenases; C575986:YfcM protein, E coli