Cloned (Comment) | Organism |
---|---|
cloning of AH gene in Escherichia coli strain JM109, expression of wild-type enzyme, active-site variants of the enzyme, and of the nitrate reductase N-terminal chaperone binding site NarG-fusion enzyme in Escherichia coli strain BL21(DE3) | Syntrophotalea acetylenica |
Crystallization (Comment) | Organism |
---|---|
purified recombinant NarG-fusion acetylene hydratase, sitting and hanging drop vapor diffusion methods, small crystals after 3 to 4 weeks, 6.5-10 mg/ml protein in solution containing 5 mM HEPES-NaOH, pH 7.5, and 7.5 mM Na2S2O4, cryoprotection by 20% v/v 2-methyl-2,4-pentanediol, X-ray diffraction structure determination and analysis | Syntrophotalea acetylenica |
Protein Variants | Comment | Organism |
---|---|---|
D13E | site-directed mutagenesis, almost inactive mutant | Syntrophotalea acetylenica |
I142A | site-directed mutagenesis, the mutant shows a marked loss of activity compared to the wild-type enzyme | Syntrophotalea acetylenica |
K48A | site-directed mutagenesis, the mutant shows unaltered activity compared to the wild-type enzyme | Syntrophotalea acetylenica |
additional information | construction of active-site variants, and of a fusion protein of the N-terminal chaperone binding site of the Escherichia coli nitrate reductase NarG to the AH gene improving the yield and activity of AH and its variants significantly, overview | Syntrophotalea acetylenica |
KM Value [mM] | KM Value Maximum [mM] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
0.014 | - |
Acetylene | pH 7.5, 30°C | Syntrophotalea acetylenica |
Localization | Comment | Organism | GeneOntology No. | Textmining |
---|---|---|---|---|
soluble | - |
Syntrophotalea acetylenica | - |
- |
Metals/Ions | Comment | Organism | Structure |
---|---|---|---|
Iron | acetylene hydratase is a tungsten, iron-sulfur enzyme, contains 4.8 mol of iron per mol of enzyme | Syntrophotalea acetylenica | |
Tungsten | acetylene hydratase is a tungsten, iron-sulfur enzyme, active W(IV) state structure, contains 0.4 mol of tungsten per mol of enzymeoverview | Syntrophotalea acetylenica |
Natural Substrates | Organism | Comment (Nat. Sub.) | Natural Products | Comment (Nat. Pro.) | Rev. | Reac. |
---|---|---|---|---|---|---|
acetylene + H2O | Syntrophotalea acetylenica | - |
acetaldehyde | - |
r | |
acetylene + H2O | Syntrophotalea acetylenica WoAcy1 / DSM 3246 | - |
acetaldehyde | - |
r |
Organism | UniProt | Comment | Textmining |
---|---|---|---|
Syntrophotalea acetylenica | - |
- |
- |
Syntrophotalea acetylenica WoAcy1 / DSM 3246 | - |
- |
- |
Reaction | Comment | Organism | Reaction ID |
---|---|---|---|
acetaldehyde = acetylene + H2O | active-site access, active-site architecture, and reaction mechanism, overview | Syntrophotalea acetylenica |
Specific Activity Minimum [µmol/min/mg] | Specific Activity Maximum [µmol/min/mg] | Comment | Organism |
---|---|---|---|
additional information | - |
activity of different purifications, overview | Syntrophotalea acetylenica |
14.2 | - |
purified recombinant enzyme, pH 7.5, 30°C | Syntrophotalea acetylenica |
Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
---|---|---|---|---|---|---|
acetylene + H2O | - |
Syntrophotalea acetylenica | acetaldehyde | - |
r | |
acetylene + H2O | the active site residues Asp13, Lys48, and Ile142 are involved in catalysis. Asp13 close to W(IV) coordinates two molybdopterin-guanosine-dinucleotide ligands, Lys48 couples the [4Fe-4S] cluster to the W site, and Ile142 is part of a hydrophobic ring at the end of the substrate access channel designed to accommodate the substrate acetylene | Syntrophotalea acetylenica | acetaldehyde | - |
r | |
acetylene + H2O | - |
Syntrophotalea acetylenica WoAcy1 / DSM 3246 | acetaldehyde | - |
r | |
acetylene + H2O | the active site residues Asp13, Lys48, and Ile142 are involved in catalysis. Asp13 close to W(IV) coordinates two molybdopterin-guanosine-dinucleotide ligands, Lys48 couples the [4Fe-4S] cluster to the W site, and Ile142 is part of a hydrophobic ring at the end of the substrate access channel designed to accommodate the substrate acetylene | Syntrophotalea acetylenica WoAcy1 / DSM 3246 | acetaldehyde | - |
r |
Subunits | Comment | Organism |
---|---|---|
More | analysis of the active W(IV) state three-dimensional structure, overview | Syntrophotalea acetylenica |
Temperature Optimum [°C] | Temperature Optimum Maximum [°C] | Comment | Organism |
---|---|---|---|
50 | - |
- |
Syntrophotalea acetylenica |
pH Optimum Minimum | pH Optimum Maximum | Comment | Organism |
---|---|---|---|
6 | 6.5 | - |
Syntrophotalea acetylenica |
Cofactor | Comment | Organism | Structure |
---|---|---|---|
molybdenum cofactor | molybdopterin cofactor, 0.94 mol per mol of enzyme in purified recombinant enzyme | Syntrophotalea acetylenica |