Crystallization (Comment) | Organism |
---|---|
wild-type Escherichia coli biotin carboxylase and mutant E296A in complex with its substrates biotin, bicarbonate, and Mg-ADP, at 2.0 A resolution. Residue Glu296 is the general base that extracts the proton from bicarbonate, and Arg338 is the residue that stabilizes the enolate biotin intermediate in the carboxylation reaction. The B domain of biotin carboxylase is positioned closer to the active site, leading to a 2-A shift in the bound position of the adenine nucleotide and bringing it near the bicarbonate for catalysis. One of the oxygenatoms of bicarbonate is located in the correct position to initiate the nucleophilic attack on ATP to form the carboxyphosphate intermediate. The phosphate group, derived from decomposition of carboxyphosphate, is the general base that extracts the proton on this N1 atom | Escherichia coli |
Protein Variants | Comment | Organism |
---|---|---|
E296A | 50fold decrease in catalytic efficiency, crystallization data | Escherichia coli |
R338A | 250fold decrease in catalytic efficiency | Escherichia coli |
KM Value [mM] | KM Value Maximum [mM] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
16.2 | - |
HCO3- | pH 8.5, wild-type | Escherichia coli | |
18.9 | - |
HCO3- | pH 8.5, mutant R338A | Escherichia coli | |
22.7 | - |
biotin | pH 7.4, mutant E296A | Escherichia coli | |
35.1 | - |
biotin | pH 7.4 wild-type | Escherichia coli | |
41.8 | - |
biotin | pH 7.4, mutant R338A | Escherichia coli | |
57.5 | - |
HCO3- | pH 8.5, mutant E296A | Escherichia coli |
Organism | UniProt | Comment | Textmining |
---|---|---|---|
Escherichia coli | P24182 | - |
- |
Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
---|---|---|---|---|---|---|
ATP + biotin + HCO3- | - |
Escherichia coli | ADP + phosphate + carboxybiotin | - |
? |
Turnover Number Minimum [1/s] | Turnover Number Maximum [1/s] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
0.0019 | - |
HCO3- | pH 8.5, mutant R338A | Escherichia coli | |
0.0025 | - |
biotin | pH 7.4, mutant R338A | Escherichia coli | |
0.028 | - |
biotin | pH 7.4, mutant E296A | Escherichia coli | |
0.034 | - |
HCO3- | pH 8.5, mutant E296A | Escherichia coli | |
0.44 | - |
HCO3- | pH 8.5, wild-type | Escherichia coli | |
0.58 | - |
biotin | pH 7.4 wild-type | Escherichia coli |
kcat/KM Value [1/mMs-1] | kcat/KM Value Maximum [1/mMs-1] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
0.00006 | - |
biotin | pH 7.4, mutant R338A | Escherichia coli | |
0.0001 | - |
HCO3- | pH 8.5, mutant R338A | Escherichia coli | |
0.000594 | - |
HCO3- | pH 8.5, mutant E296A | Escherichia coli | |
0.00123 | - |
biotin | pH 7.4, mutant E296A | Escherichia coli | |
0.0165 | - |
biotin | pH 7.4 wild-type | Escherichia coli | |
0.0272 | - |
HCO3- | pH 8.5, wild-type | Escherichia coli |