Cloned (Comment) | Organism |
---|---|
gene MTH_150, recombinant overexpression of N-terminally His6-tagged wild-type and mutant enzymes in Escherichia coli strain BL21 Gold (DE3) | Methanothermobacter thermautotrophicus |
Crystallization (Comment) | Organism |
---|---|
purified detagged recombinant wild-type and mutant enzymes, hanging drop vapour diffusion, method screening, mixing of 0.002 ml of 10 mg/ml protein in 10 mM HEPES, pH 7.5, 0.5 M NaCl, and 5 mM ligand NAD+ or NADP+, with 0.002 ml of reservoir solution containing 1.5-1.6 M ammonium sulfate, 5% glycerol, 100 mM Tris, pH 8.0, and equilibration against 0.5 ml of reservoir solution, 20°C, 48-72 h, X-ray diffraction structure determination and analysis at 1.9-2.3 A resolution, molecular replacement | Methanothermobacter thermautotrophicus |
Protein Variants | Comment | Organism |
---|---|---|
R11K | site-directed mutagenesis, like the wild-type enzyme, the enzyme mutant traps a molecule of NADP+ in the active site. This NADP+ molecule is bound in a conformation different from that displayed by NAD+ in the native enzyme complex, the mutant shows similar activity and kinetics compared to the wild-type enzyme | Methanothermobacter thermautotrophicus |
R136K | site-directed mutagenesis, like the wild-type enzyme, the enzyme mutant traps a molecule of NADP+ in the active site. This NADP+ molecule is bound in a conformation different from that displayed by NAD+ in the native enzyme complex, the mutant shows similar activity and kinetics compared to the wild-type enzyme | Methanothermobacter thermautotrophicus |
R47E | site-directed mutagenesis, like the wild-type enzyme, the enzyme mutant traps a molecule of NADP+ in the active site. This NADP+ molecule is bound in a conformation different from that displayed by NAD+ in the native enzyme complex | Methanothermobacter thermautotrophicus |
R47K | site-directed mutagenesis, like the wild-type enzyme, the enzyme mutant traps a molecule of NADP+ in the active site. This NADP+ molecule is bound in a conformation different from that displayed by NAD+ in the native enzyme complex | Methanothermobacter thermautotrophicus |
Metals/Ions | Comment | Organism | Structure |
---|---|---|---|
Mg2+ | required | Methanothermobacter thermautotrophicus |
Natural Substrates | Organism | Comment (Nat. Sub.) | Natural Products | Comment (Nat. Pro.) | Rev. | Reac. |
---|---|---|---|---|---|---|
ATP + nicotinamide ribonucleotide | Methanothermobacter thermautotrophicus | - |
diphosphate + NAD+ | - |
r | |
ATP + nicotinamide ribonucleotide | Methanothermobacter thermautotrophicus ATCC 29096 | - |
diphosphate + NAD+ | - |
r | |
additional information | Methanothermobacter thermautotrophicus | bifunctional enzyme, that also shows nicotinate-nucleotide adenylyltransferase activity, EC 2.7.7.18. The enzyme reversibly catalyzes two closely related reactions: the biosyntheses of NAD+ and its nicotinic acid analogue (NaAD+) from their respective mononucleotide precursors and ATP | ? | - |
? | |
additional information | Methanothermobacter thermautotrophicus ATCC 29096 | bifunctional enzyme, that also shows nicotinate-nucleotide adenylyltransferase activity, EC 2.7.7.18. The enzyme reversibly catalyzes two closely related reactions: the biosyntheses of NAD+ and its nicotinic acid analogue (NaAD+) from their respective mononucleotide precursors and ATP | ? | - |
? |
Organism | UniProt | Comment | Textmining |
---|---|---|---|
Methanothermobacter thermautotrophicus | O26253 | gene MTH_150 | - |
Methanothermobacter thermautotrophicus ATCC 29096 | O26253 | gene MTH_150 | - |
Purification (Comment) | Organism |
---|---|
recombinant His6-tagged wild-type and mutant enzymes from Escherichia coli strain BL21 Gold (DE3) by anion exchange and nickel affinity chromatography, tag cleavage by thrombin, dialysis, and ultrafiltration | Methanothermobacter thermautotrophicus |
Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
---|---|---|---|---|---|---|
ATP + nicotinamide ribonucleotide | - |
Methanothermobacter thermautotrophicus | diphosphate + NAD+ | - |
r | |
ATP + nicotinamide ribonucleotide | - |
Methanothermobacter thermautotrophicus ATCC 29096 | diphosphate + NAD+ | - |
r | |
additional information | bifunctional enzyme, that also shows nicotinate-nucleotide adenylyltransferase activity, EC 2.7.7.18. The enzyme reversibly catalyzes two closely related reactions: the biosyntheses of NAD+ and its nicotinic acid analogue (NaAD+) from their respective mononucleotide precursors and ATP | Methanothermobacter thermautotrophicus | ? | - |
? | |
additional information | the wild-type enzyme traps a molecule of NADP+ in the active site. This NADP+ molecule is bound in a conformation different from that displayed by NAD+ in the enzyme complex. Binding structure of wild-type and mutant enzymes with NAD+ and NADP+, detailed overview | Methanothermobacter thermautotrophicus | ? | - |
? | |
additional information | bifunctional enzyme, that also shows nicotinate-nucleotide adenylyltransferase activity, EC 2.7.7.18. The enzyme reversibly catalyzes two closely related reactions: the biosyntheses of NAD+ and its nicotinic acid analogue (NaAD+) from their respective mononucleotide precursors and ATP | Methanothermobacter thermautotrophicus ATCC 29096 | ? | - |
? | |
additional information | the wild-type enzyme traps a molecule of NADP+ in the active site. This NADP+ molecule is bound in a conformation different from that displayed by NAD+ in the enzyme complex. Binding structure of wild-type and mutant enzymes with NAD+ and NADP+, detailed overview | Methanothermobacter thermautotrophicus ATCC 29096 | ? | - |
? |
Synonyms | Comment | Organism |
---|---|---|
nicotinamide mononucleotide adenylyltransferase | - |
Methanothermobacter thermautotrophicus |
NMNAT | - |
Methanothermobacter thermautotrophicus |
Temperature Optimum [°C] | Temperature Optimum Maximum [°C] | Comment | Organism |
---|---|---|---|
65 | - |
assay at | Methanothermobacter thermautotrophicus |
pH Optimum Minimum | pH Optimum Maximum | Comment | Organism |
---|---|---|---|
7.5 | - |
assay at | Methanothermobacter thermautotrophicus |
General Information | Comment | Organism |
---|---|---|
metabolism | nicotinamide mononucleotide adenylyltransferase (NMNAT) catalyzes the biosynthesis of NAD+ and NaAD+ | Methanothermobacter thermautotrophicus |
additional information | residues Arg11 and Arg136 are implicated in binding the phosphate groups of the ATP substrate. Residue Arg47 does not interact with either NMN or ATP substrates directly, but is deemed to play a role in binding as it is proximal to Arg11 and Arg136, plasticity of the active site | Methanothermobacter thermautotrophicus |