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Literature summary extracted from

  • Kang, Y.N.; Tran, A.; White, R.H.; Ealick, S.E.
    A novel function for the N-terminal nucleophile hydrolase fold demonstrated by the structure of an archaeal inosine monophosphate cyclohydrolase (2007), Biochemistry, 46, 5050-5062.
    View publication on PubMedView publication on EuropePMC

Cloned(Commentary)

EC Number Cloned (Comment) Organism
3.5.4.10 overexpression of His-tagged enzyme in Escherichia coli strain BL21 Methanothermobacter thermautotrophicus
3.5.4.10 overexpression of wild-type and mutant enzymes in Escherichia coli strain BL21 Methanothermobacter thermautotrophicus

Crystallization (Commentary)

EC Number Crystallization (Comment) Organism
3.5.4.10 MthPurO in complex with either IMP or 5-aminoimidazole-4-carboxamide ribonucleotide, crystals from 30 mg/ml protein in 25-30% MPD, 0.2 M ammonium acetate, and 0.1 M sodium citrate buffer, pH 5.6, 0.001 ml protein solution is mixed with 0.001 ml of reservoir solution, equilibration against 0.5 mL reservoir solution, 24°C, od-shaped crystals, 3-4 weeks, X-ray diffraction structure determination and analysis at 2.0 A and 2.6 A resolution, respectively, method optimization, molecular replacement and modelling Methanothermobacter thermautotrophicus
3.5.4.10 MthPurO in complex with either IMP or 5-aminoimidazole-4-carboxamide ribonucleotide, X-ray diffraction structure determination and analysis at 2.0 A and 2.6 A resolution, respectively Methanothermobacter thermautotrophicus

Protein Variants

EC Number Protein Variants Comment Organism
3.5.4.10 C61A site-directed mutagenesis, the mutant shows 8fold increased activity compared to the wild-type enzyme Methanothermobacter thermautotrophicus
3.5.4.10 E102Q site-directed mutagenesis, inactive mutant Methanothermobacter thermautotrophicus
3.5.4.10 R31K site-directed mutagenesis, the mutant shows 76% reduced activity compared to the wild-type enzyme Methanothermobacter thermautotrophicus
3.5.4.10 Y59F site-directed mutagenesis, the mutant shows 34% reduced activity compared to the wild-type enzyme Methanothermobacter thermautotrophicus

Metals/Ions

EC Number Metals/Ions Comment Organism Structure
3.5.4.10 Mg2+
-
 Methanothermobacter thermautotrophicus

Molecular Weight [Da]

EC Number Molecular Weight [Da] Molecular Weight Maximum [Da] Comment Organism
3.5.4.10 22000
-
 x * 22000, recombinant enzyme, SDS-PAGE Methanothermobacter thermautotrophicus

Organism

EC Number Organism UniProt Comment Textmining
3.5.4.10 Methanothermobacter thermautotrophicus
-
 gene MJ0626
-
3.5.4.10 Methanothermobacter thermautotrophicus O27089 gene MTH1020
-

Purification (Commentary)

EC Number Purification (Comment) Organism
3.5.4.10 recombinant His-tagged enzyme from Escherichia coli strain BL21 by cobalt affinity chromatography, elution with imidazole, to homogeneity Methanothermobacter thermautotrophicus
3.5.4.10 soluble recombinant wild-type and mutant enzymes from Escherichia coli strain BL21 by heat treatment for 15 min at 70°C, and anion exchange chromatography, to homogeneity Methanothermobacter thermautotrophicus

Reaction

EC Number Reaction Comment Organism Reaction ID
3.5.4.10 IMP + H2O = 5-formamido-1-(5-phospho-D-ribosyl)imidazole-4-carboxamide catalytic mechanism, Glu104 is essential for catalysis Methanothermobacter thermautotrophicus
a
3.5.4.10 IMP + H2O = 5-formamido-1-(5-phospho-D-ribosyl)imidazole-4-carboxamide two possible catalytic mechanisms for cyclization, Glu102 is essential for catalysis Methanothermobacter thermautotrophicus
a

Specific Activity [micromol/min/mg]

EC Number Specific Activity Minimum [µmol/min/mg] Specific Activity Maximum [µmol/min/mg] Comment Organism
3.5.4.10 0.44
-
 purified recombinant mutant R31K Methanothermobacter thermautotrophicus
3.5.4.10 1.2
-
 purified recombinant mutant Y59F Methanothermobacter thermautotrophicus
3.5.4.10 1.8
-
 purified recombinant wild-type enzyme Methanothermobacter thermautotrophicus
3.5.4.10 32
-
 purified recombinant mutant C61A Methanothermobacter thermautotrophicus

Substrates and Products (Substrate)

EC Number Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
3.5.4.10 5-formaminoimidazole-4-carboxamide ribonucleotide the enzyme catalyzes the cyclization of 5-formaminoimidazole-4-carboxamide ribonucleotide, FAICAR, to IMP in the final step of de novo purine biosynthesis Methanothermobacter thermautotrophicus IMP + H2O
-
 ?

Subunits

EC Number Subunits Comment Organism
3.5.4.10 ? x * 22000, recombinant enzyme, SDS-PAGE Methanothermobacter thermautotrophicus
3.5.4.10 More MthPurO has a four-layered alphabetabetaalpha core structure, showing an N-terminal nucleophile hydrolase fold. The active site is located at the deep pocket between two central beta-sheets and contains residues strictly conserved within PurOs Methanothermobacter thermautotrophicus
3.5.4.10 tetramer MthPurO has a four-layered alphabetabetaalpha core structure, showing an N-terminal nucleophile hydrolase fold. The active site is located at the deep pocket between two central beta-sheets and contains residues strictly conserved within PurOs, structure analysis, modelling, detailed overview Methanothermobacter thermautotrophicus

Synonyms

EC Number Synonyms Comment Organism
3.5.4.10 inosine 5'-monophosphate cyclohydrolase
-
 Methanothermobacter thermautotrophicus
3.5.4.10 MjPurO
-
 Methanothermobacter thermautotrophicus
3.5.4.10 MthPurO
-
 Methanothermobacter thermautotrophicus
3.5.4.10 PurO
-
 Methanothermobacter thermautotrophicus

Temperature Optimum [°C]

EC Number Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
3.5.4.10 60
-
 assay at Methanothermobacter thermautotrophicus

pH Optimum

EC Number pH Optimum Minimum pH Optimum Maximum Comment Organism
3.5.4.10 7.2
-
 assay at Methanothermobacter thermautotrophicus