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Literature summary extracted from

  • Kawakami, R.; Sakuraba, H.; Goda, S.; Tsuge, H.; Ohshima, T.
    Refolding, characterization and crystal structure of (S)-malate dehydrogenase from the hyperthermophilic archaeon Aeropyrum pernix (2009), Biochim. Biophys. Acta, 1794, 1496-1504.
    View publication on PubMed

Cloned(Commentary)

EC Number Cloned (Comment) Organism
1.1.1.82 expression of MDH in Escherichia coli strain BL21(DE3) in inclusion bodies Aeropyrum pernix

Crystallization (Commentary)

EC Number Crystallization (Comment) Organism
1.1.1.82 purified refolded reconbinant enzyme, sitting drop vapor diffusion method, 0.002 ml of protein solution containing 8.9 mg/ml protein in 50 mM Tris-HCl, pH7.5, containing 0.2 M NaCl, are mixed with an equal volume of the reservoir solution, comprised of 0.1 M CHES, pH 9.5, and 40% PEG 600, and equilibrated against 0.15 ml of reservoir solution at 20°C, 2 weeks, X-ray diffraction structure determination and analysis at 2.9 A resolution and room temperature Aeropyrum pernix

KM Value [mM]

EC Number KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
1.1.1.82 0.019
-
 (S)-malate pH 10.0, 50°C, with cofactor NADP+, recombinant enzyme Aeropyrum pernix
1.1.1.82 0.12
-
 (S)-malate pH 10.0, 50°C, with cofactor NAD+, recombinant enzyme Aeropyrum pernix
1.1.1.82 0.2
-
 (2S,3R)-tartrate pH 10.0, 50°C, with cofactor NAD+, recombinant enzyme Aeropyrum pernix
1.1.1.82 1.2
-
 (2S,3S)-tartrate pH 10.0, 50°C, with cofactor NAD+, recombinant enzyme Aeropyrum pernix
1.1.1.82 2 3 (2S,3S)-tartrate pH 10.0, 50°C, with cofactor NADP+, recombinant enzyme Aeropyrum pernix
1.1.1.82 5.8
-
 (2S,3R)-tartrate pH 10.0, 50°C, with cofactor NADP+, recombinant enzyme Aeropyrum pernix

Localization

EC Number Localization Comment Organism GeneOntology No. Textmining

Molecular Weight [Da]

EC Number Molecular Weight [Da] Molecular Weight Maximum [Da] Comment Organism
1.1.1.82 33489
-
 4 * 34000, recombinant enzyme, SDS-PAGE, 4 * 33489, sequence calculation Aeropyrum pernix
1.1.1.82 34000
-
 4 * 34000, recombinant enzyme, SDS-PAGE, 4 * 33489, sequence calculation Aeropyrum pernix
1.1.1.82 110000
-
 recombinant enzyme, gel filtration Aeropyrum pernix

Organism

EC Number Organism UniProt Comment Textmining
1.1.1.82 Aeropyrum pernix Q9YEA1 a strict aerobic hyperthermophilic archaeon isolated from a coastal thermal vent in Japan
-

Purification (Commentary)

EC Number Purification (Comment) Organism
1.1.1.82 recombinant MDH from Escherichia coli inclusion bodies by solubilization, refolding, dialysis, and gel filtration Aeropyrum pernix

Renatured (Commentary)

EC Number Renatured (Comment) Organism
1.1.1.82 solubilization and refolding of recombinant MDH from Escherichia coli inclusion bodies, the enzyme is dissolved in 6 M guanidine-HCl and gradually refolded to the active enzyme through dilution of the denaturant, conditions: refolding buffer 1.5 L of 0.1 M Tris-HCl, pH 7.5, containing 2 mM EDTA and 0.4 M L-arginine, incubation for 36 h at 4°C Aeropyrum pernix

Substrates and Products (Substrate)

EC Number Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
1.1.1.82 (2S,3R)-tartrate + NAD(P)+
-
 Aeropyrum pernix ? + NAD(P)H + H+
-
 ?
1.1.1.82 (2S,3S)-tartrate + NAD(P)+
-
 Aeropyrum pernix ? + NAD(P)H + H+
-
 ?
1.1.1.82 (S)-malate + NAD(P)+
-
 Aeropyrum pernix oxaloacetate + NAD(P)H + H+
-
 ?

Subunits

EC Number Subunits Comment Organism
1.1.1.82 More structure qand sequence comaprsion, overall structure, overview Aeropyrum pernix
1.1.1.82 tetramer 4 * 34000, recombinant enzyme, SDS-PAGE, 4 * 33489, sequence calculation Aeropyrum pernix

Synonyms

EC Number Synonyms Comment Organism
1.1.1.82 (S)-malate dehydrogenase
-
 Aeropyrum pernix
1.1.1.82 MDH
-
 Aeropyrum pernix

Temperature Optimum [°C]

EC Number Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
1.1.1.82 95
-
 above Aeropyrum pernix

Temperature Range [°C]

EC Number Temperature Minimum [°C] Temperature Maximum [°C] Comment Organism
1.1.1.82 50 95
-
 Aeropyrum pernix

Temperature Stability [°C]

EC Number Temperature Stability Minimum [°C] Temperature Stability Maximum [°C] Comment Organism
1.1.1.82 50
-
 purified recombinant enzyme, 30 min, pH 5.0-10.5, stable Aeropyrum pernix
1.1.1.82 90
-
 purified recombinant enzyme, 10 min, stable. The hyperthermostability of the Aeropyrum pernix MDH is likely attributable to its smaller cavity volume and larger numbers of ion pairs and ion-pair networks, but the molecular strategy for thermostability may be specific for each enzyme Aeropyrum pernix
1.1.1.82 100
-
 purified recombinant enzyme, 10 min, stable. The hyperthermostability of the Aeropyrum pernix MDH is likely attributable to its smaller cavity volume and larger numbers of ion pairs and ion-pair networks, but the molecular strategy for thermostability may be specific for each enzyme Aeropyrum pernix

Turnover Number [1/s]

EC Number Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
1.1.1.82 0.16
-
 (2S,3R)-tartrate pH 10.0, 50°C, with cofactor NAD+, recombinant enzyme Aeropyrum pernix
1.1.1.82 0.2
-
 (2S,3R)-tartrate pH 10.0, 50°C, with cofactor NADP+, recombinant enzyme Aeropyrum pernix
1.1.1.82 0.37
-
 (2S,3S)-tartrate pH 10.0, 50°C, with cofactor NAD+, recombinant enzyme Aeropyrum pernix
1.1.1.82 1.4
-
 (S)-malate pH 10.0, 50°C, with cofactor NADP+, recombinant enzyme Aeropyrum pernix
1.1.1.82 2.6
-
 (S)-malate pH 10.0, 50°C, with cofactor NAD+, recombinant enzyme Aeropyrum pernix
1.1.1.82 4.1
-
 (2S,3S)-tartrate pH 10.0, 50°C, with cofactor NADP+, recombinant enzyme Aeropyrum pernix

pH Optimum

EC Number pH Optimum Minimum pH Optimum Maximum Comment Organism
1.1.1.82 11
-
-
 Aeropyrum pernix

pH Stability

EC Number pH Stability pH Stability Maximum Comment Organism
1.1.1.82 5 10.5 purified recombinant enzyme, 30 min, 50°C, stable Aeropyrum pernix

Cofactor

EC Number Cofactor Comment Organism Structure
1.1.1.82 NAD+ NADP+ is the preferred cofactor compared to NAD+ with substrate (S)-malate, while NAD+ is preferred with substrate tartrate. Ala at position 53 is responsible for coenzyme specificity, and the next residue, Arg, is important for NADP+ binding, structural analysis and comparison Aeropyrum pernix
1.1.1.82 NADP+ NADP+ is the preferred cofactor compared to NAD+ with substrate (S)-malate, while NAD+ is preferred with substrate tartrate. Ala at position 53 is responsible for coenzyme specificity, and the next residue, Arg, is important for NADP+ binding, structural analysis and comparison Aeropyrum pernix