TY - JOUR
T1 - Conserved Cysteine Residues of GidA Are Essential for Biogenesis of 5-Carboxymethylaminomethyluridine at tRNA Anticodon
AU - Osawa, Takuo
AU - Ito, Koichi
AU - Inanaga, Hideko
AU - Nureki, Osamu
AU - Tomita, Kozo
AU - Numata, Tomoyuki
N1 - Funding Information:
We thank T. Suzuki and M. Kimura for valuable and critical comments, and suggestions for this manuscript. We thank H. Hori for donating the A. aeolicus genomic DNA. We also thank the beamline staff at NW12A, BL-5A, and BL-17A of KEK (Ibaraki, Japan) for technical assistance during data collection. This work was supported by a PRESTO Program grant from Japan Science and Technology, a Grant-in-Aid for Young Scientists from JSPS, and grants from the Sumitomo Foundation, the Kurata Memorial Hitachi Science and Technology Foundation, and the Kato Memorial Bioscience Foundation (to T.N.).
PY - 2009/5/13
Y1 - 2009/5/13
N2 - The 5-carboxymethylaminomethyl modification of uridine (cmnm5U) at the anticodon first position occurs in tRNAs that read split codon boxes ending with purine. This modification is crucial for correct translation, by restricting codon-anticodon wobbling. Two conserved enzymes, GidA and MnmE, participate in the cmnm5U modification process. Here we determined the crystal structure of Aquifex aeolicus GidA at 2.3 Å resolution. The structure revealed the tight interaction of GidA with FAD. Structure-based mutation analyses allowed us to identify two conserved Cys residues in the vicinity of the FAD-binding site that are essential for the cmnm5U modification in vivo. Together with mutational analysis of MnmE, we propose a mechanism for the cmnm5U modification process where GidA, but not MnmE, attacks the C6 atom of uridine by a mechanism analogous to that of thymidylate synthase. We also present a tRNA-docking model that provides structural insights into the tRNA recognition mechanism for efficient modification.
AB - The 5-carboxymethylaminomethyl modification of uridine (cmnm5U) at the anticodon first position occurs in tRNAs that read split codon boxes ending with purine. This modification is crucial for correct translation, by restricting codon-anticodon wobbling. Two conserved enzymes, GidA and MnmE, participate in the cmnm5U modification process. Here we determined the crystal structure of Aquifex aeolicus GidA at 2.3 Å resolution. The structure revealed the tight interaction of GidA with FAD. Structure-based mutation analyses allowed us to identify two conserved Cys residues in the vicinity of the FAD-binding site that are essential for the cmnm5U modification in vivo. Together with mutational analysis of MnmE, we propose a mechanism for the cmnm5U modification process where GidA, but not MnmE, attacks the C6 atom of uridine by a mechanism analogous to that of thymidylate synthase. We also present a tRNA-docking model that provides structural insights into the tRNA recognition mechanism for efficient modification.
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U2 - 10.1016/j.str.2009.03.013
DO - 10.1016/j.str.2009.03.013
M3 - Article
C2 - 19446527
AN - SCOPUS:65149088650
SN - 0969-2126
VL - 17
SP - 713
EP - 724
JO - Structure with Folding & design
JF - Structure with Folding & design
IS - 5
ER -