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Literature summary for 6.1.1.27 extracted from

  • Hauenstein, S.I.; Hou, Y.M.; Perona, J.J.
    The homotetrameric phosphoseryl-tRNA synthetase from Methanosarcina mazei exhibits half-of-the-sites activity (2008), J. Biol. Chem., 283, 21997-22006.
    View publication on PubMedView publication on EuropePMC
Search for more literature for 6.1.1.27

Cloned(Commentary)

Cloned (Comment) Organism
expression of His-tagged wild-type and mutant enzymes in Escherichia coli Methanosarcina mazei

Protein Variants

Protein Variants Comment Organism
T307S mutant reveals a 3.2fold improvement in kcat/Km for phosphothreonyl adenylate synthesis, as compared with wild-type SepRS. The mutant is unable to transfer phosphothreonine to tRNACys at greater than 10% plateau levels Methanosarcina mazei
T307S site-directed mutagenesis, the mutant shows increased activity with phosphothreonine, thus reduced substrate specificity Methanosarcina mazei

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
additional information
-
 additional information recombinant enzyme, ATP/diphosphate and aminoacylation kinetics, Michaelis-Menten kinetics Methanosarcina mazei
0.04
-
 O-phospho-L-serine wild-type enzyme Methanosarcina mazei
0.15
-
 O-phospho-L-serine pH 7.5, 37°C, recombinant mutant T307S Methanosarcina mazei
0.27
-
 O-phospho-L-serine pH 7.5, 37°C, recombinant enzyme Methanosarcina mazei
0.93
-
 O-phospho-L-threonine pH 7.5, 37°C, recombinant mutant T307S Methanosarcina mazei
2.2
-
 O-phospho-L-threonine pH 7.5, 37°C, recombinant enzyme Methanosarcina mazei
8.4
-
 O-phospho-L-threonine wild-type enzyme Methanosarcina mazei

Metals/Ions

Metals/Ions Comment Organism Structure
Mg2+
-
 Methanosarcina mazei

Molecular Weight [Da]

Molecular Weight [Da] Molecular Weight Maximum [Da] Comment Organism
60909
-
 4 * 60909, calculated from sequence Methanosarcina mazei
60909
-
 4 * 60909, sequence calculation, 4 * 68992, homotetramer alpha4, mass spectrometry Methanosarcina mazei
68992
-
 4 * 60909, sequence calculation, 4 * 68992, homotetramer alpha4, mass spectrometry Methanosarcina mazei
250000
-
 gel filtration Methanosarcina mazei
250000
-
 recombinant enzyme,native PAGE, and gel filtration Methanosarcina mazei
255400
-
 recombinant enzyme, mass spectrometry Methanosarcina mazei

Natural Substrates/ Products (Substrates)

Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
ATP + O-phospho-L-serine + tRNACys Methanosarcina mazei
-
 AMP + diphosphate + O-phospho-L-seryl-tRNACys
-
 ?
ATP + O-phospho-L-serine + tRNACys Methanosarcina mazei half-of-the-sites activity: the tetrameric enzyme binds two tRNAs. Only two of the four chemically equivalent subunits catalyze formation of phosphoseryl adenylate AMP + diphosphate + O-phospho-L-seryl-tRNACys
-
 ?

Organism

Organism UniProt Comment Textmining
Methanosarcina mazei
-
-
-

Purification (Commentary)

Purification (Comment) Organism
recombinant Methanosarcina mazei
recombinant His-tagged wild-type and mutant enzymes from Escherichia coli to homogeneity by nickel affinity chromatography, the His-tag is cleaved off Methanosarcina mazei

Specific Activity [micromol/min/mg]

Specific Activity Minimum [µmol/min/mg] Specific Activity Maximum [µmol/min/mg] Comment Organism
additional information
-
 recombinant enzyme, ATP-diphosphate exchange activity Methanosarcina mazei

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
ATP + O-phospho-L-serine + tRNACys
-
 Methanosarcina mazei AMP + diphosphate + O-phospho-L-seryl-tRNACys
-
 ?
ATP + O-phospho-L-serine + tRNACys half-of-the-sites activity: the tetrameric enzyme binds two tRNAs. Only two of the four chemically equivalent subunits catalyze formation of phosphoseryl adenylate Methanosarcina mazei AMP + diphosphate + O-phospho-L-seryl-tRNACys
-
 ?
ATP + O-phospho-L-serine + tRNACys half-of-the-sites activity: the tetrameric enzyme binds two tRNAs. Only two of the four chemically equivalent subunits catalyze formation of phosphoseryl adenylate. Efficient phosphoserylation by SepRS requires methylation of tRNACys at the N1 position of G37 in the anticodon loop Methanosarcina mazei AMP + diphosphate + O-phospho-L-seryl-tRNACys
-
 ?
ATP + O-phospho-L-serine + tRNACys the tetrameric enzyme binds two tRNAs and only two of the four chemically equivalent subunits catalyze formation of phosphoseryl adenylate. tRNACys binding to SepRS also enhances the capacity of the enzyme to discriminate among amino acids, indicating the existence of functional connectivity between the tRNA and amino acid binding sites of the enzyme Methanosarcina mazei AMP + diphosphate + O-phospho-L-seryl-tRNACys
-
 ?
ATP + O-phospho-L-threonine + tRNACys low activity Methanosarcina mazei AMP + diphosphate + O-phospho-L-threonyl-tRNACys
-
 ?
ATP + O-phospho-L-threonine + tRNACys about 35% of the plateau aminoacylation observed with O-phospho-L-serine Methanosarcina mazei AMP + diphosphate + O-phospho-L-threonyl-tRNACys
-
 ?
additional information SepRS is able to discriminate against the noncognate amino acids glutamate, serine, and phosphothreonine without the need for a separate hydrolytic editing site Methanosarcina mazei ?
-
 ?
additional information SepRS is able to discriminate against the noncognate amino acids glutamate, serine, and phosphothreonine without the need for a separate hydrolytic editing site. Determination of the ATP-diphosphate exchange activity. Serine and glutamate are poor substrates, overview Methanosarcina mazei ?
-
 ?

Subunits

Subunits Comment Organism
More the tetrameric enzyme binds two tRNAs and only two of the four chemically equivalent subunits catalyze formation of phosphoseryl adenylate, active site titrations reveal calculation of 2.3 active sites per tetramer, overview Methanosarcina mazei
tetramer 4 * 60909, calculated from sequence Methanosarcina mazei
tetramer 4 * 60909, sequence calculation, 4 * 68992, homotetramer alpha4, mass spectrometry Methanosarcina mazei

Synonyms

Synonyms Comment Organism
More the enzyme is a class II tRNA synthetase Methanosarcina mazei
phosphoseryl-tRNA synthetase
-
 Methanosarcina mazei
SepRS
-
 Methanosarcina mazei

Temperature Optimum [°C]

Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
37
-
 assay at Methanosarcina mazei

Turnover Number [1/s]

Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
0.0054
-
 O-phospho-L-threonine wild-type enzyme Methanosarcina mazei
0.014
-
 O-phospho-L-threonine pH 7.5, 37°C, recombinant enzyme Methanosarcina mazei
0.021
-
 O-phospho-L-threonine pH 7.5, 37°C, recombinant mutant T307S Methanosarcina mazei
0.45
-
 O-phospho-L-serine wild-type enzyme Methanosarcina mazei
10.6
-
 O-phospho-L-serine pH 7.5, 37°C, recombinant enzyme Methanosarcina mazei
11.2
-
 O-phospho-L-serine pH 7.5, 37°C, recombinant mutant T307S Methanosarcina mazei

pH Optimum

pH Optimum Minimum pH Optimum Maximum Comment Organism
7.5
-
 assay at Methanosarcina mazei

Cofactor

Cofactor Comment Organism Structure
ATP
-
 Methanosarcina mazei