Abstract
Glutaminyl-transfer RNA (Gln-tRNA(Gln)) in archaea is synthesized in a pretranslational amidation of misacylated Glu-tRNA(Gln) by the heterodimeric Glu-tRNA(Gln) amidotransferase GatDE. Here we report the crystal structure of the Methanothermobacter thermautotrophicus GatDE complexed to tRNA(Gln) at 3.15 angstroms resolution. Biochemical analysis of GatDE and of tRNA(Gln) mutants characterized the catalytic centers for the enzyme's three reactions (glutaminase, kinase, and amidotransferase activity). A 40 angstrom-long channel for ammonia transport connects the active sites in GatD and GatE. tRNA(Gln) recognition by indirect readout based on shape complementarity of the D loop suggests an early anticodon-independent RNA-based mechanism for adding glutamine to the genetic code.
Publication types
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Research Support, N.I.H., Extramural
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Research Support, Non-U.S. Gov't
MeSH terms
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Acylation
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Adenosine Triphosphate / metabolism
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Ammonia / metabolism
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Anticodon
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Binding Sites
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Catalytic Domain
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Computer Simulation
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Crystallography, X-Ray
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Dimerization
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Genetic Code*
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Glutamine / metabolism*
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Hydrogen Bonding
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Magnesium / metabolism
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Methanobacteriaceae / enzymology*
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Methanobacteriaceae / genetics
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Models, Molecular
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Mutation
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Nitrogenous Group Transferases / chemistry*
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Nitrogenous Group Transferases / metabolism*
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Nucleic Acid Conformation
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Protein Structure, Quaternary
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Protein Structure, Secondary
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Protein Structure, Tertiary
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RNA, Archaeal / chemistry*
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RNA, Archaeal / metabolism
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RNA, Transfer, Gln / chemistry*
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RNA, Transfer, Gln / metabolism
Substances
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Anticodon
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RNA, Archaeal
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RNA, Transfer, Gln
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Glutamine
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Ammonia
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Adenosine Triphosphate
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Nitrogenous Group Transferases
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glutamyl-tRNA(Gln) amidotransferase
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Magnesium