Title | G-domain dimerization orchestrates the tRNA wobble modification reaction in the MnmE/GidA complex. |
Publication Type | Journal Article |
Year of Publication | 2009 |
Authors | Meyer, S., A. Wittinghofer, and W. Versées |
Journal | J Mol Biol |
Volume | 392 |
Issue | 4 |
Pagination | 910-22 |
Date Published | 2009 Oct 02 |
ISSN | 1089-8638 |
Keywords | Bacterial Proteins, Escherichia coli Proteins, GTP Phosphohydrolases, Guanosine Triphosphate, Humans, Hydrolysis, Models, Biological, Models, Molecular, Multiprotein Complexes, Protein Binding, Protein Multimerization, Protein Structure, Tertiary, RNA Processing, Post-Transcriptional, RNA, Transfer, Yeasts |
Abstract | MnmE and GidA are involved in the modification of wobble uridine to carboxymethylaminomethyl uridine in certain tRNAs. Malfunctioning of the human orthologs has been implicated in mitochondrial diseases. MnmE is a conserved G protein activated by dimerization. Here, we show that complex formation between MnmE and GidA involves large conformational changes that induce G-domain dimerization of MmnE and that GidA co-stimulates GTP hydrolysis on MnmE. Starting from a structural model of the complex, we identify interface mutations disrupting complex formation or communication. Although GidA does not directly contact the G-domains, conformational changes in MnmE, induced by G-domain dimerization in the triphosphate state, regulate the affinity for GidA. We developed a tRNA modification assay and demonstrate for the first time in vitro that the MnmE/GidA complex catalyzes incorporation of glycine into tRNA. An intact MnmE/GidA complex rather than their sequential action is crucial for in vitro modification. Since only GTP, but not GDP or non-hydrolyzable GTP analogs, drives the MnmE/GidA-catalyzed modification reaction, we conclude that GTP hydrolysis is essential for activity. We finally show that an active GTPase, an intact MnmE/GidA communication, and dimerization of G-domains are necessary for in vivo functioning since mutations disrupting either result in a respiratory deficient phenotype in yeast. |
DOI | 10.1016/j.jmb.2009.07.004 |
Alternate Journal | J. Mol. Biol. |
PubMed ID | 19591841 |
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