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Literature summary extracted from

  • Uechi, K.; Takata, G.; Yoneda, K.; Ohshima, T.; Sakuraba, H.
    Structure of D-tagatose 3-epimerase-like protein from Methanocaldococcus jannaschii (2014), Acta Crystallogr. Sect. F, 70, 890-895 .
    View publication on PubMedView publication on EuropePMC

Cloned(Commentary)

EC Number Cloned (Comment) Organism
5.1.3.31 gene MJ1311, recombinant expression in Escherichia coli in inclusion bodies Methanocaldococcus jannaschii

Crystallization (Commentary)

EC Number Crystallization (Comment) Organism
5.1.3.31 purified enzyme, sitting drop vapour diffusion method, mixing of 0.001 ml of 10.3 mg/ml protein in 10 mM potassium phosphate, pH 7.0, with 0.001 ml of reservoir solution composed of 0.2 M ammonium acetate, 30% isopropanol, 0.1 M Tris-HCl, pH 7.5, and equilibration against 0.1 ml reservoir solution, 3-7 days, 20°C, X-ray diffraction structure determination and analysis at 2.64 A resolution, heavy metal labeleing, single isomorphous replacement with anomalous scattering, and structure molecular modeling Methanocaldococcus jannaschii

Metals/Ions

EC Number Metals/Ions Comment Organism Structure
5.1.3.31 Mn2+ four Mn2+ ions per enzyme homodimer, structure modeling, overview Methanocaldococcus jannaschii

Molecular Weight [Da]

EC Number Molecular Weight [Da] Molecular Weight Maximum [Da] Comment Organism
5.1.3.31 60000
-
 recombinant enzyme, gel filtration Methanocaldococcus jannaschii

Organism

EC Number Organism UniProt Comment Textmining
5.1.3.31 Methanocaldococcus jannaschii Q58707
-
-
5.1.3.31 Methanocaldococcus jannaschii ATCC 43067 / 43067D / DSM 2661 / JAL-1 / JCM 10045 / NBRC 100440 Q58707
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-

Purification (Commentary)

EC Number Purification (Comment) Organism
5.1.3.31 recombinant solublized and refolded enzyme expressed in Escherichia coli by dialysis and gel filtration Methanocaldococcus jannaschii

Renatured (Commentary)

EC Number Renatured (Comment) Organism
5.1.3.31 recombinant enzyme from Escherichia coli inclusion bodies by treatment with 20 ml 10 mM Tris-HCl buffer, pH 7.5, containing 1.0 mM EDTA, 4% Triton X-100, incubation at room temperature for 30 min, twice. Then the inclusion bodies are treated with denaturant solution containing 50 mM Tris-HCl buffer, pH 7.5, containing 6 M guanidine-HCl, 0.2 M NaCl, 1 mM EDTA overnight at 4°C. The solubilized enzyme (100 mg protein in 100 ml solution) is gently dropped into 1 l of refolding buffer containing 0.1 M Tris-HCl, pH 7.5, 1 mM MnCl2, and 0.4 M L-arginine, and incubated for 16 h at 4°C, followed by ultrafiltration Methanocaldococcus jannaschii

Subunits

EC Number Subunits Comment Organism
5.1.3.31 homodimer 2 x 30000, recombinant enzyme, SDS-PAGE Methanocaldococcus jannaschii
5.1.3.31 More the asymmetric unit contained two homologous subunits, and the dimer is generated by twofold symmetry. In MJ1311p, Glu125, Leu126 and Trp127 from one subunit are located over the metal-ion-binding site of the other subunit and contribute to the active site, narrowing the substrate-binding cleft. Three-dimensional structural analysis of MJ1311p, overview. The enzyme MJ1311p monomer is folded into an (alpha/beta)8 barrel carrying four additional helical segments, alpha1', alpha2', alpha4', and alpha6', which are inserted before alpha1, alpha2, alpha4, and alpha6, respectively. The quaternary-structural arrangement of MJ1311p is notably different from those of D-TE family enzymes Methanocaldococcus jannaschii

Synonyms

EC Number Synonyms Comment Organism
5.1.3.31 MJ1311
-
 Methanocaldococcus jannaschii
5.1.3.31 MJ1311p
-
 Methanocaldococcus jannaschii

General Information

EC Number General Information Comment Organism
5.1.3.31 evolution the enzyme belongs to the D-tagatose 3-epimerase (D-TE) family enzymes. The overall fold of the subunit proves to be similar to those of the D-tagatose 3-epimerase from Pseudomonas cichorii and the D-psicose 3-epimerases from Agrobacterium tumefaciens and Clostridium cellulolyticum. But the situation at the subunit-subunit interface differs substantially from that in D-tagatose 3-epimerase family enzymes. In MJ1311p, Glu125, Leu126 and Trp127 from one subunit are located over the metal-ion-binding site of the other subunit and contribute to the active site, narrowing the substrate-binding cleft. Moreover, the nine residues comprising a trinuclear zinc centre in endonuclease IV are strictly conserved in MJ1311p, although a distinct groove involved in DNA binding is not present. The active-site architecture of MJ1311p is quite unique and is substantially different from those of D-tagatose 3-epimerase family enzymes and endonuclease IV Methanocaldococcus jannaschii