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Literature summary extracted from

  • Thamm, A.M.; Li, G.; Taja-Moreno, M.; Gerdes, S.Y.; de Crecy-Lagard, V.; Bruner, S.D.; Hanson, A.D.
    A strictly monofunctional bacterial hydroxymethylpyrimidine phosphate kinase precludes damaging errors in thiamin biosynthesis (2017), Biochem. J., 474, 2887-2895 .
    View publication on PubMed

Cloned(Commentary)

EC Number Cloned (Comment) Organism
2.7.4.7 gene thiD2, functional complementation of an Escherichia coli DELTAthiD knockout mutant strain, recombinant expression of His-tagged enzyme in Escherichia coli strain BL21-CodonPlus (DE3)-RIPL Campylobacter concisus
2.7.4.7 gene thiD2, functional complementation of an Escherichia coli DELTAthiD knockout mutant strain, recombinant expression of His-tagged enzyme in Escherichia coli strain BL21-CodonPlus (DE3)-RIPL Sulfurimonas denitrificans

Protein Variants

EC Number Protein Variants Comment Organism
2.7.1.49 additional information construction of a thiD knockout mutant strain, complementation of the Escherichia coli DELTA thiD knockout mutant is possible by heterologous expression of gene ThiD2, ThiD2 proteins catalyze phosphorylation of HMP monophosphate, but not of HMP or its toxic analogues and damage products such as bacimethrin and 5-(hydroxymethyl)-2-methylpyrimidin-4-ol. As strictly monofunctional HMP monophosphate kinases (EC 2.7.4.7), ThiD2 proteins eliminate a potentially fatal vulnerability of canonical ThiD, at the cost of the ability to reclaim HMP formed by thiamin turnover Escherichia coli
2.7.4.7 additional information construction of a thiD knockout mutant strain, complementation of the Escherichia coli DELTA thiD knockout mutant is possible by heterologous expression of gene ThiD2, ThiD2 proteins catalyze phosphorylation of HMP monophosphate, but not of HMP or its toxic analogues and damage products such as bacimethrin and 5-(hydroxymethyl)-2-methylpyrimidin-4-ol. As strictly monofunctional HMP monophosphate kinases (EC 2.7.4.7), ThiD2 proteins eliminate a potentially fatal vulnerability of canonical ThiD, at the cost of the ability to reclaim HMP formed by thiamin turnover Escherichia coli
2.7.4.7 additional information complementation of the Escherichia coli DELTA thiD knockout mutant is possible by heterologous expression of gene ThiD2. Strains expressing a standalone ThiD2 protein are 10fold more resistant than wild-type against bacimethrin toxicity. ThiD2 genes confer resistance to bacimethrin Campylobacter concisus
2.7.4.7 additional information complementation of the Escherichia coli DELTA thiD knockout mutant is possible by heterologous expression of gene ThiD2. Strains expressing a standalone ThiD2 protein are 10fold more resistant than wild-type against bacimethrin toxicity. ThiD2 genes confer resistance to bacimethrin Sulfurimonas denitrificans

KM Value [mM]

EC Number KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
2.7.1.49 0.00104
-
 4-Amino-5-hydroxymethyl-2-methylpyrimidine pH 7.5, temperature not specified in the publication, recombinant enzyme ThiD Escherichia coli
2.7.4.7 0.00232
-
 4-amino-2-methyl-5-(phosphooxymethyl)pyrimidine pH 7.5, temperature not specified in the publication, recombinant enzyme ThiD Escherichia coli
2.7.4.7 0.00512
-
 4-amino-2-methyl-5-(phosphooxymethyl)pyrimidine pH 7.5, temperature not specified in the publication, recombinant enzyme ThiD2 Sulfurimonas denitrificans

Metals/Ions

EC Number Metals/Ions Comment Organism Structure
2.7.1.49 Mg2+ required Escherichia coli
2.7.4.7 Mg2+ required Campylobacter concisus
2.7.4.7 Mg2+ required Escherichia coli
2.7.4.7 Mg2+ required Sulfurimonas denitrificans

Natural Substrates/ Products (Substrates)

EC Number Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
2.7.1.49 ATP + 4-amino-2-methyl-5-(phosphooxymethyl)pyrimidine Escherichia coli
-
 ADP + 4-amino-2-methyl-5-(diphosphooxymethyl)pyrimidine
-
 ?
2.7.1.49 ATP + 4-amino-2-methyl-5-(phosphooxymethyl)pyrimidine Escherichia coli BW25113
-
 ADP + 4-amino-2-methyl-5-(diphosphooxymethyl)pyrimidine
-
 ?
2.7.1.49 ATP + 4-amino-5-hydroxymethyl-2-methylpyrimidine Escherichia coli
-
 ADP + 4-amino-2-methyl-5-(phosphooxymethyl)pyrimidine
-
 ?
2.7.1.49 ATP + 4-amino-5-hydroxymethyl-2-methylpyrimidine Escherichia coli BW25113
-
 ADP + 4-amino-2-methyl-5-(phosphooxymethyl)pyrimidine
-
 ?
2.7.4.7 ATP + 4-amino-2-methyl-5-(phosphooxymethyl)pyrimidine Escherichia coli
-
 ADP + 4-amino-2-methyl-5-(diphosphooxymethyl)pyrimidine
-
 ?
2.7.4.7 ATP + 4-amino-2-methyl-5-(phosphooxymethyl)pyrimidine Campylobacter concisus
-
 ADP + 4-amino-2-methyl-5-(diphosphooxymethyl)pyrimidine
-
 ?
2.7.4.7 ATP + 4-amino-2-methyl-5-(phosphooxymethyl)pyrimidine Sulfurimonas denitrificans
-
 ADP + 4-amino-2-methyl-5-(diphosphooxymethyl)pyrimidine
-
 ?
2.7.4.7 ATP + 4-amino-2-methyl-5-(phosphooxymethyl)pyrimidine Escherichia coli BW25113
-
 ADP + 4-amino-2-methyl-5-(diphosphooxymethyl)pyrimidine
-
 ?
2.7.4.7 ATP + 4-amino-2-methyl-5-(phosphooxymethyl)pyrimidine Campylobacter concisus 13826
-
 ADP + 4-amino-2-methyl-5-(diphosphooxymethyl)pyrimidine
-
 ?
2.7.4.7 ATP + 4-amino-2-methyl-5-(phosphooxymethyl)pyrimidine Sulfurimonas denitrificans ATCC 33889
-
 ADP + 4-amino-2-methyl-5-(diphosphooxymethyl)pyrimidine
-
 ?
2.7.4.7 ATP + 4-amino-2-methyl-5-(phosphooxymethyl)pyrimidine Sulfurimonas denitrificans DSM 1251
-
 ADP + 4-amino-2-methyl-5-(diphosphooxymethyl)pyrimidine
-
 ?

Organism

EC Number Organism UniProt Comment Textmining
2.7.1.49 Escherichia coli P76422
-
-
2.7.1.49 Escherichia coli BW25113 P76422
-
-
2.7.4.7 Campylobacter concisus A7ZG21
-
-
2.7.4.7 Campylobacter concisus 13826 A7ZG21
-
-
2.7.4.7 Escherichia coli P76422
-
-
2.7.4.7 Escherichia coli BW25113 P76422
-
-
2.7.4.7 Sulfurimonas denitrificans Q30NV2 i.e. Thiomicrospira denitrificans strain ATCC 33889 / DSM 1251
-
2.7.4.7 Sulfurimonas denitrificans ATCC 33889 Q30NV2 i.e. Thiomicrospira denitrificans strain ATCC 33889 / DSM 1251
-
2.7.4.7 Sulfurimonas denitrificans DSM 1251 Q30NV2 i.e. Thiomicrospira denitrificans strain ATCC 33889 / DSM 1251
-

Purification (Commentary)

EC Number Purification (Comment) Organism
2.7.4.7 recombinant His-tagged enzyme from Escherichia coli strain BL21-CodonPlus (DE3)-RIPL by nickel affinity chromatography and desalting gel filtration Campylobacter concisus
2.7.4.7 recombinant His-tagged enzyme from Escherichia coli strain BL21-CodonPlus (DE3)-RIPL by nickel affinity chromatography and desalting gel filtration Sulfurimonas denitrificans

Substrates and Products (Substrate)

EC Number Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
2.7.1.49 ATP + 4-amino-2-methyl-5-(phosphooxymethyl)pyrimidine
-
 Escherichia coli ADP + 4-amino-2-methyl-5-(diphosphooxymethyl)pyrimidine
-
 ?
2.7.1.49 ATP + 4-amino-2-methyl-5-(phosphooxymethyl)pyrimidine reaction of EC 2.7.4.7 Escherichia coli ADP + 4-amino-2-methyl-5-(diphosphooxymethyl)pyrimidine
-
 ?
2.7.1.49 ATP + 4-amino-2-methyl-5-(phosphooxymethyl)pyrimidine
-
 Escherichia coli BW25113 ADP + 4-amino-2-methyl-5-(diphosphooxymethyl)pyrimidine
-
 ?
2.7.1.49 ATP + 4-amino-2-methyl-5-(phosphooxymethyl)pyrimidine reaction of EC 2.7.4.7 Escherichia coli BW25113 ADP + 4-amino-2-methyl-5-(diphosphooxymethyl)pyrimidine
-
 ?
2.7.1.49 ATP + 4-amino-5-hydroxymethyl-2-methylpyrimidine
-
 Escherichia coli ADP + 4-amino-2-methyl-5-(phosphooxymethyl)pyrimidine
-
 ?
2.7.1.49 ATP + 4-amino-5-hydroxymethyl-2-methylpyrimidine
-
 Escherichia coli BW25113 ADP + 4-amino-2-methyl-5-(phosphooxymethyl)pyrimidine
-
 ?
2.7.1.49 additional information the enzyme performs double phosphorylation on 4-amino-5-hydroxymethyl-2-methylpyrimidine (HMP) Escherichia coli ?
-
 ?
2.7.1.49 additional information the enzyme performs double phosphorylation on 4-amino-5-hydroxymethyl-2-methylpyrimidine (HMP) Escherichia coli BW25113 ?
-
 ?
2.7.4.7 ATP + 4-amino-2-methyl-5-(phosphooxymethyl)pyrimidine
-
 Escherichia coli ADP + 4-amino-2-methyl-5-(diphosphooxymethyl)pyrimidine
-
 ?
2.7.4.7 ATP + 4-amino-2-methyl-5-(phosphooxymethyl)pyrimidine
-
 Campylobacter concisus ADP + 4-amino-2-methyl-5-(diphosphooxymethyl)pyrimidine
-
 ?
2.7.4.7 ATP + 4-amino-2-methyl-5-(phosphooxymethyl)pyrimidine
-
 Sulfurimonas denitrificans ADP + 4-amino-2-methyl-5-(diphosphooxymethyl)pyrimidine
-
 ?
2.7.4.7 ATP + 4-amino-2-methyl-5-(phosphooxymethyl)pyrimidine
-
 Escherichia coli BW25113 ADP + 4-amino-2-methyl-5-(diphosphooxymethyl)pyrimidine
-
 ?
2.7.4.7 ATP + 4-amino-2-methyl-5-(phosphooxymethyl)pyrimidine
-
 Campylobacter concisus 13826 ADP + 4-amino-2-methyl-5-(diphosphooxymethyl)pyrimidine
-
 ?
2.7.4.7 ATP + 4-amino-2-methyl-5-(phosphooxymethyl)pyrimidine
-
 Sulfurimonas denitrificans ATCC 33889 ADP + 4-amino-2-methyl-5-(diphosphooxymethyl)pyrimidine
-
 ?
2.7.4.7 ATP + 4-amino-2-methyl-5-(phosphooxymethyl)pyrimidine
-
 Sulfurimonas denitrificans DSM 1251 ADP + 4-amino-2-methyl-5-(diphosphooxymethyl)pyrimidine
-
 ?
2.7.4.7 additional information the enzyme performs double phosphorylation on 4-amino-5-hydroxymethyl-2-methylpyrimidine (HMP) Escherichia coli ?
-
 ?
2.7.4.7 additional information the enzyme performs double phosphorylation on 4-amino-5-hydroxymethyl-2-methylpyrimidine (HMP) Campylobacter concisus ?
-
 ?
2.7.4.7 additional information the enzyme performs double phosphorylation on 4-amino-5-hydroxymethyl-2-methylpyrimidine (HMP) Sulfurimonas denitrificans ?
-
 ?
2.7.4.7 additional information the enzyme performs double phosphorylation on 4-amino-5-hydroxymethyl-2-methylpyrimidine (HMP) Campylobacter concisus 13826 ?
-
 ?
2.7.4.7 additional information the enzyme performs double phosphorylation on 4-amino-5-hydroxymethyl-2-methylpyrimidine (HMP) Sulfurimonas denitrificans ATCC 33889 ?
-
 ?
2.7.4.7 additional information the enzyme performs double phosphorylation on 4-amino-5-hydroxymethyl-2-methylpyrimidine (HMP) Sulfurimonas denitrificans DSM 1251 ?
-
 ?

Synonyms

EC Number Synonyms Comment Organism
2.7.1.49 HMP kinase
-
 Escherichia coli
2.7.1.49 hydroxymethylpyrimidine phosphate kinase
-
 Escherichia coli
2.7.1.49 additional information see also EC 2.7.4.7 Escherichia coli
2.7.1.49 ThiD
-
 Escherichia coli
2.7.4.7 CCC13826_0338 locus name Campylobacter concisus
2.7.4.7 CcThiD2
-
 Campylobacter concisus
2.7.4.7 HMP-P kinase
-
 Escherichia coli
2.7.4.7 HMP-P kinase
-
 Campylobacter concisus
2.7.4.7 HMP-P kinase
-
 Sulfurimonas denitrificans
2.7.4.7 hydroxymethylpyrimidine phosphate kinase
-
 Escherichia coli
2.7.4.7 hydroxymethylpyrimidine phosphate kinase
-
 Campylobacter concisus
2.7.4.7 hydroxymethylpyrimidine phosphate kinase
-
 Sulfurimonas denitrificans
2.7.4.7 additional information see also EC 2.7.1.49 Escherichia coli
2.7.4.7 Suden_2055 locus name Sulfurimonas denitrificans
2.7.4.7 TdThiD2
-
 Sulfurimonas denitrificans
2.7.4.7 thiamine phosphate synthase UniProt Campylobacter concisus
2.7.4.7 ThiD
-
 Escherichia coli
2.7.4.7 ThiD2
-
 Campylobacter concisus
2.7.4.7 ThiD2
-
 Sulfurimonas denitrificans
2.7.4.7 ThiD2 domain-containing protein UniProt Sulfurimonas denitrificans

Turnover Number [1/s]

EC Number Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
2.7.1.49 0.285
-
 4-Amino-5-hydroxymethyl-2-methylpyrimidine pH 7.5, temperature not specified in the publication, recombinant enzyme ThiD Escherichia coli
2.7.4.7 0.011
-
 4-amino-2-methyl-5-(phosphooxymethyl)pyrimidine pH 7.5, temperature not specified in the publication, recombinant enzyme ThiD2 Sulfurimonas denitrificans
2.7.4.7 0.034
-
 4-amino-2-methyl-5-(phosphooxymethyl)pyrimidine pH 7.5, temperature not specified in the publication, recombinant enzyme ThiD Escherichia coli

pH Optimum

EC Number pH Optimum Minimum pH Optimum Maximum Comment Organism
2.7.1.49 7.5
-
 assay at Escherichia coli
2.7.4.7 7.5
-
 assay at Escherichia coli
2.7.4.7 7.5
-
 assay at Campylobacter concisus
2.7.4.7 7.5
-
 assay at Sulfurimonas denitrificans

General Information

EC Number General Information Comment Organism
2.7.1.49 evolution ThiD is a member of the ribokinase family, but differs from other members in catalyzing two consecutive phosphorylations. The other members of the family catalyze only the phosphorylation of a hydroxymethyl group to give a monophosphate, i.e. the equivalent of the HMP kinase reaction. The HMP kinase activity of ThiD is hence presumably ancestral and the HMP-P kinase activity is an evolutionary novelty Escherichia coli
2.7.1.49 metabolism the enzyme takes part in the bacterial thiamin biosynthesis and salvage pathways, overview Escherichia coli
2.7.1.49 physiological function the bifunctional canonical kinase (ThiD) that converts the thiamin biosynthesis intermediate hydroxymethylpyrimidine (HMP) monophosphate into the diphosphate (EC 2.7.4.7) can also very efficiently convert free HMP into the monophosphate (EC 2.7.1.49) in prokaryotes, plants, and fungi. This HMP kinase activity enables salvage of HMP, but it is not substrate-specific and so allows toxic HMP analogues and damage products to infiltrate the thiamin biosynthesis pathway Escherichia coli
2.7.4.7 evolution ThiD is a member of the ribokinase family, but differs from other members in catalyzing two consecutive phosphorylations. The other members of the family catalyze only the phosphorylation of a hydroxymethyl group to give a monophosphate, i.e. the equivalent of the HMP kinase reaction. The HMP kinase activity of ThiD is hence presumably ancestral and the HMP-P kinase activity is an evolutionary novelty Escherichia coli
2.7.4.7 metabolism the enzyme takes part in the bacterial thiamin biosynthesis and salvage pathways, overview Escherichia coli
2.7.4.7 metabolism the enzyme takes part in the bacterial thiamin biosynthesis and salvage pathways, overview Campylobacter concisus
2.7.4.7 metabolism the enzyme takes part in the bacterial thiamin biosynthesis and salvage pathways, overview Sulfurimonas denitrificans
2.7.4.7 physiological function the bifunctional canonical kinase (ThiD) that converts the thiamin biosynthesis intermediate hydroxymethylpyrimidine (HMP) monophosphate into the diphosphate (EC 2.7.4.7) can also very efficiently convert free HMP into the monophosphate (EC 2.7.1.49) in prokaryotes, plants, and fungi. This HMP kinase activity enables salvage of HMP, but it is not substrate-specific and so allows toxic HMP analogues and damage products to infiltrate the thiamin biosynthesis pathway Escherichia coli
2.7.4.7 physiological function ThiD2 proteins catalyze phosphorylation of HMP monophosphate, but not of HMP or its toxic analogues and damage products such as bacimethrin and 5-(hydroxymethyl)-2-methylpyrimidin-4-ol. As strictly monofunctional HMP monophosphate kinases (EC 2.7.4.7), ThiD2 proteins eliminate a potentially fatal vulnerability of canonical ThiD, at the cost of the ability to reclaim HMP formed by thiamin turnover. ThiD2 genes confer resistance to bacimethrin Campylobacter concisus
2.7.4.7 physiological function ThiD2 proteins catalyze phosphorylation of HMP monophosphate, but not of HMP or its toxic analogues and damage products such as bacimethrin and 5-(hydroxymethyl)-2-methylpyrimidin-4-ol. As strictly monofunctional HMP monophosphate kinases (EC 2.7.4.7), ThiD2 proteins eliminate a potentially fatal vulnerability of canonical ThiD, at the cost of the ability to reclaim HMP formed by thiamin turnover. ThiD2 genes confer resistance to bacimethrin Sulfurimonas denitrificans

kcat/KM [mM/s]

EC Number kcat/KM Value [1/mMs-1] kcat/KM Value Maximum [1/mMs-1] Substrate Comment Organism Structure
2.7.1.49 274.04
-
 4-Amino-5-hydroxymethyl-2-methylpyrimidine pH 7.5, temperature not specified in the publication, recombinant enzyme ThiD Escherichia coli
2.7.4.7 2.15
-
 4-amino-2-methyl-5-(phosphooxymethyl)pyrimidine pH 7.5, temperature not specified in the publication, recombinant enzyme ThiD2 Sulfurimonas denitrificans
2.7.4.7 14.66
-
 4-amino-2-methyl-5-(phosphooxymethyl)pyrimidine pH 7.5, temperature not specified in the publication, recombinant enzyme ThiD Escherichia coli