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Literature summary extracted from

  • Herring, S.; Ambrogelly, A.; Gundllapalli, S.; O'Donoghue, P.; Polycarpo, C.R.; Soll, D.
    The amino-terminal domain of pyrrolysyl-tRNA synthetase is dispensable in vitro but required for in vivo activity (2007), FEBS Lett., 581, 3197-3203.
    View publication on PubMedView publication on EuropePMC

Cloned(Commentary)

EC Number Cloned (Comment) Organism
6.1.1.26 gene pylS, DNA and amino acid sequence determination and analysis, expression in Escherichia coli BL21(DE3) as His-tagged enzyme Methanosarcina thermophila
6.1.1.26 gene pylS, expression in Escherichia coli BL21(DE3) as His-tagged enzyme Desulfitobacterium hafniense
6.1.1.26 gene pylS, expression of His-tagged wild-type and mutant enzymes in Escherichia coli BL21(DE3) Methanosarcina barkeri

Protein Variants

EC Number Protein Variants Comment Organism
6.1.1.26 D2A site-directed mutagenesis, the mutant shows 95% reduced activity compared to the wild-type enzyme Methanosarcina barkeri
6.1.1.26 D2A/K4A site-directed mutagenesis, the mutant shows almost completely reduced activity compared to the wild-type enzyme Methanosarcina barkeri
6.1.1.26 D7A site-directed mutagenesis, the mutant shows only slightly reduced activity compared to the wild-type enzyme Methanosarcina barkeri
6.1.1.26 G21L site-directed mutagenesis, the mutant shows only slightly reduced activity compared to the wild-type enzyme Methanosarcina barkeri
6.1.1.26 H24A site-directed mutagenesis, the mutant shows 95% reduced activity compared to the wild-type enzyme Methanosarcina barkeri
6.1.1.26 I26G site-directed mutagenesis, the mutant shows 80% reduced activity compared to the wild-type enzyme Methanosarcina barkeri
6.1.1.26 K3A site-directed mutagenesis, the mutant shows 80% reduced activity compared to the wild-type enzyme Methanosarcina barkeri
6.1.1.26 K4A site-directed mutagenesis, the mutant shows 95% reduced activity compared to the wild-type enzyme Methanosarcina barkeri
6.1.1.26 additional information construction of several truncation mutants, which show 90-99% reduced activity compared to the wild-type enzyme, overview Methanosarcina barkeri
6.1.1.26 R19A site-directed mutagenesis, the mutant shows only slightly reduced activity compared to the wild-type enzyme Methanosarcina barkeri
6.1.1.26 S11A site-directed mutagenesis, the mutant shows 95% reduced activity compared to the wild-type enzyme Methanosarcina barkeri
6.1.1.26 S11A/T13A site-directed mutagenesis, the mutant shows almost completely reduced activity compared to the wild-type enzyme Methanosarcina barkeri
6.1.1.26 S18A site-directed mutagenesis, the mutant shows only slightly reduced activity compared to the wild-type enzyme Methanosarcina barkeri
6.1.1.26 T13A site-directed mutagenesis, the mutant shows 95% reduced activity compared to the wild-type enzyme Methanosarcina barkeri
6.1.1.26 W16A site-directed mutagenesis, the mutant shows 35% reduced activity compared to the wild-type enzyme Methanosarcina barkeri

KM Value [mM]

EC Number KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
6.1.1.26 0.00015 0.00098 additional information equilibrium binding analysis, dissociation constants of the enzyme with tRNAPyl from different species, overview Methanosarcina barkeri
6.1.1.26 0.00016 0.001 additional information equilibrium binding analysis, dissociation constants of the enzyme with tRNAPyl from different species, overview Methanosarcina thermophila
6.1.1.26 0.0053 0.0069 additional information equilibrium binding analysis, dissociation constants of the enzyme with tRNAPyl from different species, overview Desulfitobacterium hafniense

Metals/Ions

EC Number Metals/Ions Comment Organism Structure
6.1.1.26 Mg2+
-
 Desulfitobacterium hafniense

Natural Substrates/ Products (Substrates)

EC Number Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
6.1.1.26 ATP + L-pyrrolysine + tRNAPyl Methanosarcina barkeri
-
 AMP + diphosphate + L-pyrrolysyl-tRNAPyl
-
 ?
6.1.1.26 ATP + L-pyrrolysine + tRNAPyl Desulfitobacterium hafniense
-
 AMP + diphosphate + L-pyrrolysyl-tRNAPyl
-
 ?
6.1.1.26 ATP + L-pyrrolysine + tRNAPyl Methanosarcina thermophila
-
 AMP + diphosphate + L-pyrrolysyl-tRNAPyl
-
 ?
6.1.1.26 additional information Methanosarcina thermophila the amino-terminal extension present in archaeal PylRSs is dispensable for in vitro activity, but required for PylRS function in vivo ?
-
 ?

Organism

EC Number Organism UniProt Comment Textmining
6.1.1.26 Desulfitobacterium hafniense
-
 a strictly anaerobic bacterium
-
6.1.1.26 Methanosarcina barkeri
-
 strains Fusaro and MS, gene pylS
-
6.1.1.26 Methanosarcina thermophila Q1L6A3 gene pylS
-

Purification (Commentary)

EC Number Purification (Comment) Organism
6.1.1.26 recombinant His-tagged enzyme from Escherichia coli BL21(DE3) Desulfitobacterium hafniense
6.1.1.26 recombinant His-tagged enzyme from Escherichia coli BL21(DE3) Methanosarcina thermophila
6.1.1.26 recombinant His-tagged wild-type and mutant enzymes from Escherichia coli BL21(DE3) Methanosarcina barkeri

Substrates and Products (Substrate)

EC Number Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
6.1.1.26 ATP + L-pyrrolysine + tRNAPyl
-
 Methanosarcina barkeri AMP + diphosphate + L-pyrrolysyl-tRNAPyl
-
 ?
6.1.1.26 ATP + L-pyrrolysine + tRNAPyl
-
 Desulfitobacterium hafniense AMP + diphosphate + L-pyrrolysyl-tRNAPyl
-
 ?
6.1.1.26 ATP + L-pyrrolysine + tRNAPyl
-
 Methanosarcina thermophila AMP + diphosphate + L-pyrrolysyl-tRNAPyl
-
 ?
6.1.1.26 ATP + L-pyrrolysine + tRNAPyl the archaeal enzyme does not distinguish between archaeal and bacterial tRNAPyl species, substrate specificity, overview Methanosarcina thermophila AMP + diphosphate + L-pyrrolysyl-tRNAPyl
-
 ?
6.1.1.26 ATP + L-pyrrolysine + tRNAPyl the archaeal enzyme does not distinguish between archaeal and bacterial tRNAPyl species, substrate specificity, overview, residues from the PylRS amino-terminal domain affect activity in vivo Methanosarcina barkeri AMP + diphosphate + L-pyrrolysyl-tRNAPyl
-
 ?
6.1.1.26 ATP + L-pyrrolysine + tRNAPyl the bacterial PylRS displays a clear preference for the homologous cognate tRNA, substrate specificity, overview Desulfitobacterium hafniense AMP + diphosphate + L-pyrrolysyl-tRNAPyl
-
 ?
6.1.1.26 additional information the amino-terminal extension present in archaeal PylRSs is dispensable for in vitro activity, but required for PylRS function in vivo Methanosarcina thermophila ?
-
 ?

Subunits

EC Number Subunits Comment Organism
6.1.1.26 More the PylRS sequence can be subdivided into three regions: the highly conserved class II aaRS catalytic core domain at the carboxy-terminal, the unique amino-terminal domain, and a highly variable region linking these two domains Desulfitobacterium hafniense
6.1.1.26 More the PylRS sequence can be subdivided into three regions: the highly conserved class II aaRS catalytic core domain at the carboxy-terminal, the unique amino-terminal domain, and a highly variable region linking these two domains, residues from the PylRS amino-terminal domain affect activity in vivo Methanosarcina barkeri
6.1.1.26 More the PylRS sequence can be subdivided into three regions: the highly conserved class II aaRS catalytic core domain at the carboxy-terminal, the unique amino-terminal domain, and a highly variable region linking these two domains, the amino-terminal extension present in archaeal PylRSs is dispensable for in vitro activity, but required for PylRS function in vivo Methanosarcina thermophila

Synonyms

EC Number Synonyms Comment Organism
6.1.1.26 PylRS
-
 Methanosarcina barkeri
6.1.1.26 PylRS
-
 Desulfitobacterium hafniense
6.1.1.26 PylRS
-
 Methanosarcina thermophila
6.1.1.26 pyrrolysyl-tRNA synthetase
-
 Methanosarcina barkeri
6.1.1.26 pyrrolysyl-tRNA synthetase
-
 Desulfitobacterium hafniense
6.1.1.26 pyrrolysyl-tRNA synthetase
-
 Methanosarcina thermophila

Temperature Optimum [°C]

EC Number Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
6.1.1.26 37
-
 assay at Methanosarcina barkeri
6.1.1.26 37
-
 assay at Desulfitobacterium hafniense
6.1.1.26 37
-
 assay at Methanosarcina thermophila

pH Optimum

EC Number pH Optimum Minimum pH Optimum Maximum Comment Organism
6.1.1.26 7.2
-
 assay at Methanosarcina barkeri
6.1.1.26 7.2
-
 assay at Desulfitobacterium hafniense
6.1.1.26 7.2
-
 assay at Methanosarcina thermophila

Cofactor

EC Number Cofactor Comment Organism Structure
6.1.1.26 ATP
-
 Methanosarcina barkeri
6.1.1.26 ATP
-
 Desulfitobacterium hafniense
6.1.1.26 ATP
-
 Methanosarcina thermophila