Cloned (Comment) | Organism |
---|---|
expressed in Escherichia coli BL21(DE3) cells | Thermotoga maritima |
MtDH is expressed in Escherichia coli BL21(DE3) | Limosilactobacillus reuteri |
the TM0298 gene subcloned into the NdeI and XhoI sites of pET24a(+) to yield plasmid pTmMtDH, from which MtDH is expressed with a C-terminal His6-tag in Escherichia coli BL21(DE3) | Thermotoga maritima |
Inhibitors | Comment | Organism | Structure |
---|---|---|---|
EDTA | activity of MtDH pretreated with 10 mM EDTA for 20 min at 37°C decreases 96%. Adding 20 mM ZnCl2 to the EDTA-treated enzyme restores activity up to 80% of the control (enzyme not treated with EDTA). Adding 20 mM CoCl2 and 20 mM MnCl2 to the EDTA-treated enzyme restores activity up to 132% and 94% of the control, respectively. In contrast, adding 20 mM MgCl2 or CaCl2 does not increase the activity of the EDTA-treated enzyme.; activity of MtDH pretreated with 10 mM for 20 min at 37°C decreases 96%. Adding 20 mM ZnCl2 to the EDTA-treated enzyme restores activity up to 80% of the control. Adding 20 mM CoCl2 and 20 mM MnCl2 to the EDTA-treated enzyme restores activity up to 132% and 94% of the control, respectively | Thermotoga maritima | |
mannitol | - |
Thermotoga maritima |
KM Value [mM] | KM Value Maximum [mM] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
0.037 | - |
NADH | at 60°C and pH 6.1 | Thermotoga maritima | |
0.037 | - |
NADH | at 60°C, pH 6.1 | Thermotoga maritima | |
0.048 | - |
NADH | at 80°C and pH 6.1 | Thermotoga maritima | |
0.048 | - |
NADH | at 80°C, pH 6.1 | Thermotoga maritima | |
0.14 | - |
NAD+ | at 80°C and pH 8.3 | Thermotoga maritima | |
0.14 | - |
NAD+ | at 80°C, pH 8.3 | Thermotoga maritima | |
0.17 | - |
NADPH | at 80°C and pH 6.1 | Thermotoga maritima | |
0.17 | - |
NADPH | at 80°C, pH 6.1 | Thermotoga maritima | |
5.51 | - |
D-mannitol | at 80°C and pH 8.3 | Thermotoga maritima | |
5.51 | - |
D-mannitol | at 80°C, pH 8.3 | Thermotoga maritima | |
7.5 | - |
NADP+ | at 80°C and pH 8.3 | Thermotoga maritima | |
7.5 | - |
NADP+ | at 80°C, pH 8.3 | Thermotoga maritima | |
13.23 | - |
D-mannitol | at 60°C and pH 6.1 | Thermotoga maritima | |
13.23 | - |
D-mannitol | at 60°C, pH 6.1 | Thermotoga maritima | |
50 | - |
D-fructose | at 60°C and pH 6.1 | Thermotoga maritima | |
50 | - |
D-fructose | at 60°C, pH 6.1 | Thermotoga maritima | |
50.97 | - |
D-fructose | at 80°C and pH 6.1 | Thermotoga maritima | |
50.97 | - |
D-fructose | at 80°C, pH 6.1 | Thermotoga maritima |
Metals/Ions | Comment | Organism | Structure |
---|---|---|---|
Co2+ | - |
Thermotoga maritima | |
Mn2+ | - |
Thermotoga maritima | |
additional information | 20 mM MgCl2 or CaCl2 do not increase the activity of the EDTA-treated enzyme | Thermotoga maritima | |
Zn2+ | contains 0.69 mol of Zn2+ per subunit of enzyme | Thermotoga maritima | |
Zn2+ | MtDH contains a single catalytic zinc per subunit | Thermotoga maritima |
Molecular Weight [Da] | Molecular Weight Maximum [Da] | Comment | Organism |
---|---|---|---|
34000 | - |
4 * 34000, SDS-PAGE | Thermotoga maritima |
120000 | - |
ultracentrifugation | Thermotoga maritima |
120000 | 143000 | gel filtration or analytical ultracentrifugation | Thermotoga maritima |
120000 | 134000 | gel filtration | Thermotoga maritima |
Organism | UniProt | Comment | Textmining |
---|---|---|---|
Limosilactobacillus reuteri | Q6ECH5 | - |
- |
Thermotoga maritima | Q9WYD4 | - |
- |
Thermotoga maritima MSB8 / DSM 3109 / ATCC 43589 | Q9WYD4 | - |
- |
Purification (Comment) | Organism |
---|---|
by heat treatment at 85°C and on Ni-NTA column | Thermotoga maritima |
Ni-NTA agarose column chromatography | Thermotoga maritima |
Specific Activity Minimum [µmol/min/mg] | Specific Activity Maximum [µmol/min/mg] | Comment | Organism |
---|---|---|---|
54 | - |
purified enzyme at 80°C with D-fructose as the substrate and NADH as the cofactor | Thermotoga maritima |
54 | - |
purified enzyme, at 80°C with fructose as the substrate and NADH as the cofactor | Thermotoga maritima |
Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
---|---|---|---|---|---|---|
D-fructose + NADPH + H+ | also active on fructose with NADPH | Thermotoga maritima | D-mannitol + NADP+ | - |
? | |
D-fructose + NADPH + H+ | also active on fructose with NADPH | Thermotoga maritima MSB8 / DSM 3109 / ATCC 43589 | D-mannitol + NADP+ | - |
? | |
D-mannitol + NAD(P)+ | - |
Thermotoga maritima | D-fructose + NAD(P)H + H+ | - |
r | |
D-mannitol + NAD(P)+ | - |
Thermotoga maritima MSB8 / DSM 3109 / ATCC 43589 | D-fructose + NAD(P)H + H+ | - |
r | |
D-mannitol + NAD+ | 100% activity | Thermotoga maritima | D-fructose + NADH + H+ | - |
r | |
D-mannitol + NAD+ | 100% activity | Thermotoga maritima MSB8 / DSM 3109 / ATCC 43589 | D-fructose + NADH + H+ | - |
r | |
D-tagatose + NAD(P)+ | 29% relative activity compared to D-fructose | Thermotoga maritima | ? + NAD(P)H | - |
? | |
D-tagatose + NAD+ | 29% relative activity on D-tagatose compared to 100% activity on D-fructose | Thermotoga maritima | ? | - |
? | |
D-xylulose + NAD(P)+ | 18% relative activity compared to D-fructose | Thermotoga maritima | ? + NAD(P)H | - |
? | |
D-xylulose + NAD+ | 18% relative activity on D-xylulose compared to 100% activity on D-fructose | Thermotoga maritima | ? | - |
? | |
L-sorbose + NAD(P)+ | 5% relative activity compared to D-fructose | Thermotoga maritima | ? + NAD(P)H | - |
? | |
L-sorbose + NADH + H+ | 5% relative activity on L-sorbose compared to 100% activity on D-fructose | Thermotoga maritima | L-sorbitol + NAD+ | - |
? | |
additional information | mannitol can be produced directly from glucose in a two-step enzymatic process, using a Thermotoga neapolitana xylose isomerase mutant and TmMtDH at 60°C. No activity with glucose, xylose, threonine, arabinose, acetaldehyde, 2-butanone, sorbitol, xylitol, ethanol, or 2-butanol | Thermotoga maritima | ? | - |
? | |
additional information | TM0298 shows no detectable activity on glucose, xylose, threonine, arabinose, acetaldehyde, 2-butanone, sorbitol, xylitol, ethanol, or 2-butanol | Thermotoga maritima | ? | - |
? | |
additional information | mannitol can be produced directly from glucose in a two-step enzymatic process, using a Thermotoga neapolitana xylose isomerase mutant and TmMtDH at 60°C. No activity with glucose, xylose, threonine, arabinose, acetaldehyde, 2-butanone, sorbitol, xylitol, ethanol, or 2-butanol | Thermotoga maritima MSB8 / DSM 3109 / ATCC 43589 | ? | - |
? | |
additional information | TM0298 shows no detectable activity on glucose, xylose, threonine, arabinose, acetaldehyde, 2-butanone, sorbitol, xylitol, ethanol, or 2-butanol | Thermotoga maritima MSB8 / DSM 3109 / ATCC 43589 | ? | - |
? |
Subunits | Comment | Organism |
---|---|---|
dimer | gel filtration at room temperature, MtDH in solution | Limosilactobacillus reuteri |
octamer | gel filtration at room temperature, MtDH in solution | Limosilactobacillus reuteri |
tetramer | 4 * 34000, SDS-PAGE | Thermotoga maritima |
tetramer | gel filtration at room temperature, MtDH in solution | Limosilactobacillus reuteri |
tetramer | gel filtration or analytical ultracentrifugation, MtDH in solution | Thermotoga maritima |
Synonyms | Comment | Organism |
---|---|---|
MtDH | - |
Thermotoga maritima |
MtDH | - |
Limosilactobacillus reuteri |
TM0298 | - |
Thermotoga maritima |
Temperature Optimum [°C] | Temperature Optimum Maximum [°C] | Comment | Organism |
---|---|---|---|
90 | 100 | most active around 95°C | Thermotoga maritima |
95 | - |
- |
Thermotoga maritima |
Temperature Minimum [°C] | Temperature Maximum [°C] | Comment | Organism |
---|---|---|---|
90 | 120 | MtDH retains 63% of its activity at 120°C but shows no detectable activity at 25°C | Thermotoga maritima |
95 | 120 | retains 63% of its activity at 120°C but shows no detectable activity at room temperature | Thermotoga maritima |
Temperature Stability Minimum [°C] | Temperature Stability Maximum [°C] | Comment | Organism |
---|---|---|---|
80 | - |
half-life: 57 min | Thermotoga maritima |
80 | - |
half-lives of 57 min | Thermotoga maritima |
95 | - |
half-life: 6 min | Thermotoga maritima |
95 | - |
half-lives of 6 min | Thermotoga maritima |
pH Optimum Minimum | pH Optimum Maximum | Comment | Organism |
---|---|---|---|
additional information | - |
pH 5.5-6 is the optimum for D-fructose reduction and pH 8.3-8.6 is the optimum for mannitol reduction | Thermotoga maritima |
5.5 | 6 | optimally reduces L-fructose at pH values between pH 5.5 and 6.0 | Thermotoga maritima |
8.3 | 8.6 | optimally oxidizes L-mannitol between pH 8.3 and 8.6 | Thermotoga maritima |
pH Stability | pH Stability Maximum | Comment | Organism |
---|---|---|---|
8.5 | - |
TM0298 is sensitive to proteolytic degradation in purification protocols at pH 8.5 | Thermotoga maritima |
Cofactor | Comment | Organism | Structure |
---|---|---|---|
NAD(P)+ | its catalytic efficiency is 33times higher with NAD+ than with NADP+ | Thermotoga maritima | |
NAD(P)H | MtDH has a higher Vmax with NADPH than with NADH, whereas its catalytic efficiency is 2.2times higher with NADH than with NADPH. Cofactor specificity is due to the high density of negatively charged residues (Glu193, Asp195, and Glu196) downstream of the NAD(P) interaction site, the glycine motif | Thermotoga maritima | |
NAD+ | although MtDH has a higher Vmax with NADPH than with NADH, its catalytic efficiency is 33 times higher with NAD+ than with NADP+ | Thermotoga maritima | |
NADH | although MtDH has a higher Vmax with NADPH than with NADH, its catalytic efficiency is 2.2times higher with NADH than with NADPH | Thermotoga maritima |
Ki Value [mM] | Ki Value maximum [mM] | Inhibitor | Comment | Organism | Structure |
---|---|---|---|---|---|
99 | - |
mannitol | - |
Thermotoga maritima |