Cloned (Comment) | Organism |
---|---|
into a TOPO vector (pSGX3), and transformed into Escherichia coli TOP10 competent cells. Gene in the pSGX3 vector transformed into Escherichia coli strains XL1-Blue for transformation and BL21(DE3) for expression | Oceanobacillus iheyensis |
Protein Variants | Comment | Organism |
---|---|---|
H45Q | has no detectable activity | Oceanobacillus iheyensis |
R162N | retains a small amount of activity | Oceanobacillus iheyensis |
Y164F | has no detectable activity | Oceanobacillus iheyensis |
Y90F | is catalytically impaired. Structure of the mutant in complex with Mg2+ and galactarate has a well-defined C-terminal segment through residue 387, well-ordered electron density for galactarate, and Mg2+ ions in both metal sites for both protomers comprising the asymmetric unit | Oceanobacillus iheyensis |
KM Value [mM] | KM Value Maximum [mM] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
0.62 | - |
galactarate | wild-type, in 50 mM Hepes, pH 8.0, 5 mM MgCl2 | Oceanobacillus iheyensis | |
2 | - |
galactarate | mutant Y90F, in 50 mM Hepes, pH 8.0, 5 mM MgCl2 | Oceanobacillus iheyensis | |
4.4 | - |
galactarate | mutant H45Q, in 50 mM Hepes, pH 8.0, 5 mM MgCl2 | Oceanobacillus iheyensis |
Metals/Ions | Comment | Organism | Structure |
---|---|---|---|
Mg2+ | structure contains two Mg2+ ions located 10.4 A from one another, with one located in the canonical position in the (beta/alpha)7beta-barrel domain, the second is located in a site within the capping domain | Oceanobacillus iheyensis |
Organism | UniProt | Comment | Textmining |
---|---|---|---|
Oceanobacillus iheyensis | - |
- |
- |
Purification (Comment) | Organism |
---|---|
by centrifugation, sonication, on Ni-NTA column and by gel filtration | Oceanobacillus iheyensis |
Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
---|---|---|---|---|---|---|
galactarate | - |
Oceanobacillus iheyensis | 5-dehydro-4-deoxy D-glucarate + H2O | - |
? |
Synonyms | Comment | Organism |
---|---|---|
galactarate dehydratase | - |
Oceanobacillus iheyensis |
GalrD | - |
Oceanobacillus iheyensis |
Turnover Number Minimum [1/s] | Turnover Number Maximum [1/s] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
0.0013 | - |
galactarate | mutant H45Q, in 50 mM Hepes, pH 8.0, 5 mM MgCl2 | Oceanobacillus iheyensis | |
0.0061 | - |
galactarate | mutant Y90F, in 50 mM Hepes, pH 8.0, 5 mM MgCl2 | Oceanobacillus iheyensis | |
6.8 | - |
galactarate | wild-type, in 50 mM Hepes, pH 8.0, 5 mM MgCl2 | Oceanobacillus iheyensis |
kcat/KM Value [1/mMs-1] | kcat/KM Value Maximum [1/mMs-1] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
0.00029 | - |
galactarate | mutant H45Q, in 50 mM Hepes, pH 8.0, 5 mM MgCl2 | Oceanobacillus iheyensis | |
0.00065 | - |
galactarate | mutant R162N, in 50 mM Hepes, pH 8.0, 5 mM MgCl2 | Oceanobacillus iheyensis | |
0.0031 | - |
galactarate | mutant Y90F, in 50 mM Hepes, pH 8.0, 5 mM MgCl2 | Oceanobacillus iheyensis | |
11 | - |
galactarate | wild-type, in 50 mM Hepes, pH 8.0, 5 mM MgCl2 | Oceanobacillus iheyensis |