Cloned (Comment) | Organism |
---|---|
expression of wild-type Cel6B and mutant enzymes in Escherichia coli strain BL21(DE3) | Thermobifida fusca |
Protein Variants | Comment | Organism |
---|---|---|
D226A | site-directed mutagenesis, the mutant shows altered substrate specificity compared to the wild-type enzyme. The D226A enzyme has a very low activity on insoluble cellulose, not improved by sodium azide | Thermobifida fusca |
D226A/S232A | site-directed mutagenesis, the mutant is almost inactive and shows slightly decreased ligand binding compared to the wild-type enzyme | Thermobifida fusca |
D274A | site-directed mutagenesis, the mutant shows altered and highly reduced substrate specificity compared to the wild-type enzyme | Thermobifida fusca |
D497A | site-directed mutagenesis, the mutant shows altered substrate specificity compared to the wild-type enzyme | Thermobifida fusca |
E495A | site-directed mutagenesis, the mutant shows altered substrate specificity compared to the wild-type enzyme | Thermobifida fusca |
S232A | site-directed mutagenesis, the mutant shows altered substrate specificity compared to the wild-type enzyme. The S232A enzyme retains near wild-type activity on most substrates, but carboxymethylcellulose activity is drastically reduced | Thermobifida fusca |
Y220A | site-directed mutagenesis, the mutant shows altered substrate specificity compared to the wild-type enzyme, alomost inactive mutant | Thermobifida fusca |
KM Value [mM] | KM Value Maximum [mM] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
0.0015 | - |
2,4-dinitrophenyl cellobiose | pH 5.5, 50°C, recombinant mutant D274A | Thermobifida fusca | |
0.0023 | - |
2,4-dinitrophenyl cellobiose | pH 5.5, 50°C, recombinant wild-type enzyme | Thermobifida fusca | |
0.0065 | - |
2,4-dinitrophenyl cellobiose | pH 5.5, 50°C, recombinant mutant D226A | Thermobifida fusca | |
0.044 | - |
2,4-dinitrophenyl cellobiose | pH 5.5, 50°C, recombinant mutant S232A | Thermobifida fusca | |
0.161 | - |
2,4-dinitrophenyl cellobiose | pH 5.5, 50°C, recombinant mutant Y220A | Thermobifida fusca | |
0.214 | - |
2,4-dinitrophenyl cellobiose | pH 5.5, 50°C, recombinant mutant D497A | Thermobifida fusca |
Natural Substrates | Organism | Comment (Nat. Sub.) | Natural Products | Comment (Nat. Pro.) | Rev. | Reac. |
---|---|---|---|---|---|---|
cellulose + H2O | Thermobifida fusca | - |
cellobiose + ? | - |
? |
Organism | UniProt | Comment | Textmining |
---|---|---|---|
Thermobifida fusca | - |
- |
- |
Purification (Comment) | Organism |
---|---|
recombinant wild-type Cel6B and mutant enzymes from Escherichia coli strain BL21(DE3) by hydrophobic interaction and anion exchange chromatography | Thermobifida fusca |
Reaction | Comment | Organism | Reaction ID |
---|---|---|---|
2 cellohexaose + 2 H2O = 2 cellotriose + cellotetraose + cellobiose | exocellulase Cel6B acts by an inverting hydrolysis mechanism, that involves a proton-transferring network to carry out the catalytic base function | Thermobifida fusca |
Specific Activity Minimum [µmol/min/mg] | Specific Activity Maximum [µmol/min/mg] | Comment | Organism |
---|---|---|---|
additional information | - |
cellulose substrate type specificity of wild-type and mutant enzymes, overview | Thermobifida fusca |
Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
---|---|---|---|---|---|---|
2,4-dinitrophenyl cellobiose + H2O | - |
Thermobifida fusca | 2,4-dinitrophenol + cellobiose | - |
? | |
4-methylumbelliferyl beta-cellobioside + H2O | substrate binding analysis of wild-type enzyme and mutant D274A, mutant D274A binds 4-methylumbelliferyl beta-cellobioside to approximately the same extent as the wild-type, overview | Thermobifida fusca | 4-methylumbelliferol + cellobiose | - |
? | |
carboxymethylcellulose + H2O | - |
Thermobifida fusca | cellobiose + ? | - |
? | |
cellulose + H2O | - |
Thermobifida fusca | cellobiose + ? | - |
? | |
cellulose + H2O | activity with bacterial microcrystalline cellulose, phosphoric acid-treated swollen cellulose, phosphoric acid-treated cotton, and carboxymethyl cellulose | Thermobifida fusca | cellobiose + ? | - |
? |
Subunits | Comment | Organism |
---|---|---|
More | structural model of the Cel6B catalytic domain, structure comparisons to other exocellulases, overview | Thermobifida fusca |
Synonyms | Comment | Organism |
---|---|---|
Cel6B | - |
Thermobifida fusca |
exocellulase | - |
Thermobifida fusca |
Temperature Optimum [°C] | Temperature Optimum Maximum [°C] | Comment | Organism |
---|---|---|---|
50 | - |
assay at | Thermobifida fusca |
Turnover Number Minimum [1/s] | Turnover Number Maximum [1/s] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
0.0001 | - |
2,4-dinitrophenyl cellobiose | pH 5.5, 50°C, recombinant mutant D497A | Thermobifida fusca | |
0.0005 | - |
2,4-dinitrophenyl cellobiose | pH 5.5, 50°C, recombinant mutant S232A | Thermobifida fusca | |
0.0015 | - |
2,4-dinitrophenyl cellobiose | pH 5.5, 50°C, recombinant mutant Y220A | Thermobifida fusca | |
0.0018 | - |
2,4-dinitrophenyl cellobiose | pH 5.5, 50°C, recombinant mutant D226A | Thermobifida fusca | |
0.038 | - |
2,4-dinitrophenyl cellobiose | pH 5.5, 50°C, recombinant mutant D274A | Thermobifida fusca | |
0.057 | - |
2,4-dinitrophenyl cellobiose | pH 5.5, 50°C, recombinant wild-type enzyme | Thermobifida fusca |
pH Optimum Minimum | pH Optimum Maximum | Comment | Organism |
---|---|---|---|
5.5 | - |
assay at | Thermobifida fusca |
kcat/KM Value [1/mMs-1] | kcat/KM Value Maximum [1/mMs-1] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
additional information | - |
additional information | - |
Thermobifida fusca |