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6.1.1.17: glutamate-tRNA ligase

This is an abbreviated version!
For detailed information about glutamate-tRNA ligase, go to the full flat file.

Word Map on EC 6.1.1.17

Reaction

ATP
+
L-glutamate
+
tRNAGlu
=
AMP
+
diphosphate
+
L-glutamyl-tRNAGlu

Synonyms

bifunctional aminoacyl-tRNA synthetase, bifunctional glutamate/proline-tRNA ligase, D-GluRS, discriminating GluRS, EC 2.7.2.13, Ec-GluRS, EPRS, ERS, Ers2, GltX, GluRS, GluRS1, GluRS2, GluRSAt, Glutamate--tRNA ligase, Glutamate-tRNA synthetase, Glutamic acid translase, Glutamic acid tRNA ligase, Glutaminyl-tRNA synthetase, Glutamyl tRNA synthetase, glutamyl-prolyl tRNA synthetase, glutamyl-prolyl-tRNA synthetase, glutamyl-Q tRNAASp synthetase, Glutamyl-transfer ribonucleate synthetase, Glutamyl-transfer ribonucleic acid synthetase, Glutamyl-transfer RNA synthetase, Glutamyl-tRNA synthetase, glutamyl-tRNAsynthetase, GtS, More, P85, surface-exposed glutamyl tRNA synthetase, TM1351, tRNA modifying enzyme, YadB

ECTree

     6 Ligases
         6.1 Forming carbon-oxygen bonds
             6.1.1 Ligases forming aminoacyl-tRNA and related compounds
                6.1.1.17 glutamate-tRNA ligase

Crystallization

Crystallization on EC 6.1.1.17 - glutamate-tRNA ligase

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CRYSTALLIZATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
purified Gly-GluRS K236E/E328A, hanging drop vapour diffusion method, mixing of 0.001 ml of 20 mg/ml protein in 20 mM HEPES, pH 7.5, 50 mM NaCl, 10 mM 2-mercaptoethanol, and 50 mM ZnCl2, with 0.0012 ml of crystallization solution containing 0.1 M MOPS/HEPES-Na, pH 7.7, 0.02 M each of L-Glu.Na, DL-Ala, DL-Lys-HCl, Gly and DL-Ser, 14% w/v PEG 8000, 22% v/v ethylene glycol, 2 weeks, method optimization, X-ray diffraction structure determination and analysis at 3.5 A resolution, molecular replacement
-
purified recombinant GluRS-N-Arc1p-N complex, hanging drop vapour diffusion method, 0.002 ml protein solution containing 15 mg/ml protein in 20 mM HEPES, 150 mM NaCl, 5 mM MgCl2, 1 mM DTT, pH 7.2 with NaOH, is mixed with 0.002 ml reservoir solution containing 30-35% PEG 3350, 300-500 mM NaSCN, X-ray diffraction structure determination and analysis at 2.05 A resolution
purified recombinant truncated enzyme, 0.002 ml of 20 mg/ml protein in 20 mM HEPES, 150 mM NaCl, 5 mM MgCl2, 1 mM DTT, pH 7.2 with NaOH, mixed with 0.002 ml reservoir solution at 20°C, equilibration against 0.075 ml reservoir solution, containing 1.7-1.8 M (NH4)2SO4, 200 mM KSCN for selenomethionine-substituted crystals and 1.8-1.9 M (NH4)2SO4, 200 mM NaI for native crystals, dispersion with selenomethionine, X-ray diffraction structure determination and analysis at 2.5 A resolution, modeling
architectures of class-defining and specific domains
-
crystallization of complexes: 1. GluRS and L-Glu, 2. GluRS, tRNAGlu, and L-Glu, 3. GluRS, tRNAGlu, ATP, and L-glutamol, 4. GluRS, tRNAGlu, and L-glutamyl-sulfamoyl adenosine, by hanging drop vapour diffusion method, 5.0 mg/ml enzyme in 10 mM MOPS-Na buffer, pH 6.5, MgCl2, 5 mM 2-mercaptoethanol, 1% PEG 6000, and 2 mM L-glutamate, equilibration against a 1 ml reservoir solution containing 10% PEG at 4°C, ERS/tRNA/Glu and ERS/tRNA/ESA crystals are prepared by diffusing 1 mM L-glutamate and 0.5 mM glutamyl-sulfamoyl adenosine, i.e. ESA, respectively, into the ERS/tRNA binary complex crystals, ERS/tRNA/ATP/Eol crystals are obtained by adding both 1 mM ATP and 1 mM L-glutamol, i.e. Eol, to drops containing the ERS/tRNA binary complex, X-ray diffraction structure determination and analysis at 1.98 A, 2.4 A, 2.2 A, and 2.69 A resolution, respectively
crystallization of the enzyme in different complexes: 1. non-productively complexed with ATP and L-glutamate, 2. with ATP, 3. with tRNAGlu and ATP, 4. with tRNAGlu and the glutamyl-AMP analogue glutamol-AMP, hanging-drop method, 0.008 ml of 5.0 mg/ml protein in 10 mM Na-MOPS, pH 6.5, 5 mM MgCl2, 2.5 mM 2-mercaptoethanol, 1% PEG 6000, 1-2 mM ATP and/or 2 mM glutamate and/or 0.5 mM glutamol-AMP, plus 1 ml reservoir solution containing 10% PEG 6000 at 4 or 20°C, 3 days or more, X-ray diffraction structure determination at 1.8 A resolution, molecular replacement, and analysis
molecular modeling, internal pKa calculations, and molecular dynamics simulations for consideration of distinct, mechanistically relevant post-transfer states with charged tRNA bound to glutamyl-tRNA synthetase. The transfer of amino acid to tRNA is accompanied by the protonation of AMP to H-AMP. Subsequent migration of proton to water reduces the stability of the complex and loosens the interface both in the presence and in the absence of AMP. The subsequent undocking of AMP or tRNA then proceeds along thermodynamically competitive pathways. Release of the tRNA acceptor stem is further accelerated by the deprotonation of the alpha-ammonium group on the charging amino acid. The proposed general base is Glu41
purified recombinant enzyme in complex with tRNAGlu, hanging-drop vapour diffusion method, precipitant solution contains 37 mM Na-MOPS, pH 6.7, 22% PEG 1500, 37 mM ammonium sulfate, 1% 2-methyl-2,4-pentanediol, 10 mM MgCl2, 5 mM 2-mercaptoethanol, X-ray diffraction structure determination at 2.4 A resolution, and analysis