Cloned (Comment) | Organism |
---|---|
expressed in Escherichia coli strain HMS174-DE3, transformation with mutant plasmids, pD567N, pE572Q, pD579N, or pE841Q for mutant generation | Bacillus sp. (in: Bacteria) |
Crystallization (Comment) | Organism |
---|---|
native and selenomethionine-derivaties, 1.9 A resolution with a final R-factor of 18.2%, hanging-drop method, data-collection and statistics indicated, overall structure shown, quaternary structure, catalytic domain and active cleft determined, complex with the reaction product rhamnose determined and refined at 2.1 A with a final R-factor of 19.5% | Bacillus sp. (in: Bacteria) |
Protein Variants | Comment | Organism |
---|---|---|
D567N | mutagenesis of catalytic amino acid residues identifed by crystallization studies, enzyme activity reduced | Bacillus sp. (in: Bacteria) |
D579N | mutagenesis of catalytic amino acid residues identifed by crystallization studies, enzyme activity reduced | Bacillus sp. (in: Bacteria) |
E572Q | mutagenesis of catalytic amino acid residues identifed by crystallization studies, enzyme activity reduced | Bacillus sp. (in: Bacteria) |
E841Q | mutagenesis of catalytic amino acid residues identifed by crystallization studies, enzyme activity reduced | Bacillus sp. (in: Bacteria) |
Inhibitors | Comment | Organism | Structure |
---|---|---|---|
L-rhamnose | crystallized complex with interacting site determined, for kinetic assay different concentrations ranging between 0 and 100 mM of rhamnose used | Bacillus sp. (in: Bacteria) |
KM Value [mM] | KM Value Maximum [mM] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
additional information | - |
additional information | 2.5 micrograms per ml of purified protein used for the wild-type enzyme and 500 micrograms per ml for mutant enzymes | Bacillus sp. (in: Bacteria) | |
0.28 | - |
p-nitrophenyl-alpha-L-rhamnopyranoside | wild-type | Bacillus sp. (in: Bacteria) | |
0.3 | - |
p-nitrophenyl-alpha-L-rhamnopyranoside | E572Q | Bacillus sp. (in: Bacteria) | |
0.31 | - |
p-nitrophenyl-alpha-L-rhamnopyranoside | E841Q | Bacillus sp. (in: Bacteria) | |
0.36 | - |
p-nitrophenyl-alpha-L-rhamnopyranoside | D567N | Bacillus sp. (in: Bacteria) | |
0.48 | - |
p-nitrophenyl-alpha-L-rhamnopyranoside | D579N | Bacillus sp. (in: Bacteria) |
Metals/Ions | Comment | Organism | Structure |
---|---|---|---|
Ca2+ | calcium ions essential for crystallization, calcium binding shown in crystallized form | Bacillus sp. (in: Bacteria) |
Organism | UniProt | Comment | Textmining |
---|---|---|---|
Bacillus sp. (in: Bacteria) | Q93RE7 | rhamnosidase B, RhaB | - |
Bacillus sp. (in: Bacteria) GL1 | Q93RE7 | rhamnosidase B, RhaB | - |
Purification (Comment) | Organism |
---|---|
native and recombinant proteins, gel filtration, SDS-PAGE | Bacillus sp. (in: Bacteria) |
Specific Activity Minimum [µmol/min/mg] | Specific Activity Maximum [µmol/min/mg] | Comment | Organism |
---|---|---|---|
additional information | - |
assayed in a reaction system containing 0.4 mM p-nitrophenyl-alpha-L-rhamnopyranoside as a substrate and 50 mM potassium phosphate buffer, resultant p-nitrophenol concentration determined spectrophotometrically | Bacillus sp. (in: Bacteria) |
Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
---|---|---|---|---|---|---|
gellan + H2O | release of rhamnose from the disaccharide, hydrolytic reaction | Bacillus sp. (in: Bacteria) | alpha-L-rhamnose + ? | residues interacting with rhamnose that are crucial for enzyme catalysis and/or substrate binding identified by crystallization | ? | |
gellan + H2O | release of rhamnose from the disaccharide, hydrolytic reaction | Bacillus sp. (in: Bacteria) GL1 | alpha-L-rhamnose + ? | residues interacting with rhamnose that are crucial for enzyme catalysis and/or substrate binding identified by crystallization | ? | |
naringin + H2O | bitter flavour source in fruit | Bacillus sp. (in: Bacteria) | 4',5,7-trihydroxyflavanone-7-beta-D-glucoside + alpha-L-rhamnose | - |
? | |
naringin + H2O | bitter flavour source in fruit | Bacillus sp. (in: Bacteria) GL1 | 4',5,7-trihydroxyflavanone-7-beta-D-glucoside + alpha-L-rhamnose | - |
? | |
p-nitrophenyl-alpha-L-rhamnopyranoside + H2O | assay reaction system to determine activity of wild-type and mutant enzymes | Bacillus sp. (in: Bacteria) | p-nitrophenol + alpha-L-rhamnopyranose | - |
? | |
p-nitrophenyl-alpha-L-rhamnopyranoside + H2O | assay reaction system to determine activity of wild-type and mutant enzymes | Bacillus sp. (in: Bacteria) GL1 | p-nitrophenol + alpha-L-rhamnopyranose | - |
? |
Subunits | Comment | Organism |
---|---|---|
homodimer | crystallized form, 1908 amino acids of residues 3-956, 43 glycerol molecules, four calcium ions, 1755 water molecules identified in the crystallized form | Bacillus sp. (in: Bacteria) |
Synonyms | Comment | Organism |
---|---|---|
alpha-L-rhamnosidase | - |
Bacillus sp. (in: Bacteria) |
Temperature Optimum [°C] | Temperature Optimum Maximum [°C] | Comment | Organism |
---|---|---|---|
30 | - |
assay at | Bacillus sp. (in: Bacteria) |
pH Optimum Minimum | pH Optimum Maximum | Comment | Organism |
---|---|---|---|
7 | - |
assay at | Bacillus sp. (in: Bacteria) |
Ki Value [mM] | Ki Value maximum [mM] | Inhibitor | Comment | Organism | Structure |
---|---|---|---|---|---|
1.8 | - |
L-rhamnose | crystallized complex with interacting site analyzed | Bacillus sp. (in: Bacteria) |