EC Number | Cloned (Comment) | Organism |
---|---|---|
2.5.1.17 | expressed in Escherichia coli as an N-terminal His-tagged fusion protein | Limosilactobacillus reuteri |
EC Number | Protein Variants | Comment | Organism |
---|---|---|---|
2.5.1.17 | DELTAS183 | Km (mM) (Co+ assay): 0.0031 (ATP), 0.00007 (cob(I)alamin), kcat (1/sec) (Co+ assay): 0.023 (ATP), 0.02 (cob(I)alamin), Km (mM) (Co2+ assay): 0.0099 (ATP), 0.0163 (cob(II)alamin), kcat (1/sec) (Co+ assay): 0.017 (ATP), 0.018 (cob(II)alamin) | Limosilactobacillus reuteri |
2.5.1.17 | F112A | Km (mM) (Co+ assay): 0.0346 (ATP), 0.0019 (cob(I)alamin), kcat (1/sec) (Co+ assay): 0.0056 (ATP), 0.006 (cob(I)alamin). Mutant F112A shows no activity in the Co2+ assay. The crystal structure of mutant F112A reveal that cob(II)alamin binds to the active site as a five-coordinate species with 5,6-dimethylbenzimidazole serving as a fifth axial ligand. In the Co+ assay [adenosylation of cob(I)alamin], mutant F112A is catalytically competent and displays only a slight decrease in kcat, supposing that in the absence of a four-coordinate cob(II)alamin species, the enzyme is inactive | Limosilactobacillus reuteri |
2.5.1.17 | F112H | Km (mM) (Co+ assay): 0.0026 (ATP), 0.0028 (cob(I)alamin), kcat (1/sec) (Co+ assay): 0.00038 (ATP), 0.0042 (cob(I)alamin), Km (mM) (Co2+ assay): 0.01 (ATP), 0.053 (cob(II)alamin), kcat (1/sec) (Co+ assay): 0.00059 (ATP), 0.00067 (cob(II)alamin). The crystal structure of an mutantF112H variant show a 5,6-dimethylbenzimidazole-off/His-on interaction between the corrinoid and the enzyme, whose catalytic efficiency is 4 orders of magnitude lower than that of the wild-type protein. The analysis of the kinetic parameters of mutantF112H suggests that the F112H substitution negatively impacts product release | Limosilactobacillus reuteri |
2.5.1.17 | F112W | Km (mM) (Co+ assay): 0.002 (ATP), 0.00077 (cob(I)alamin), kcat (1/sec) (Co+ assay): 0.0018 (ATP), 0.002 (cob(I)alamin), Km (mM) (Co2+ assay): 0.0073 (ATP), 0.028 (cob(II)alamin), kcat (1/sec) (Co+ assay): 0.0028 (ATP), 0.0032 (cob(II)alamin) | Limosilactobacillus reuteri |
2.5.1.17 | F112Y | Km (mM) (Co+ assay): 0.0012 (ATP), 0.00055 (cob(I)alamin), kcat (1/sec) (Co+ assay): 0.0016 (ATP), 0.0017 (cob(I)alamin), Km (mM) (Co2+ assay): 0.0094 (ATP), 0.034 (cob(II)alamin), kcat (1/sec) (Co+ assay): 0.0023 (ATP), 0.0027 (cob(II)alamin) | Limosilactobacillus reuteri |
2.5.1.17 | F163A | Km (mM) (Co+ assay): 0.0159 (ATP), 0.00019 (cob(I)alamin), kcat (1/sec) (Co+ assay): 0.015 (ATP), 0.015 (cob(I)alamin), Km (mM) (Co2+ assay): 0.096 (ATP), 0.134 (cob(II)alamin), kcat (1/sec) (Co+ assay): 0.015 (ATP), 0.027 (cob(II)alamin) | Limosilactobacillus reuteri |
2.5.1.17 | F187A | Km (mM) (Co+ assay): 0.0012 (ATP), 0.00011 (cob(I)alamin), kcat (1/sec) (Co+ assay): 0.021 (ATP), 0.018 (cob(I)alamin), Km (mM) (Co2+ assay): 0.0079 (ATP), 0.0158 (cob(II)alamin), kcat (1/sec) (Co+ assay): 0.014 (ATP), 0.015 (cob(II)alamin) | Limosilactobacillus reuteri |
EC Number | KM Value [mM] | KM Value Maximum [mM] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|---|
2.5.1.17 | 0.00007 | - |
cob(I)alamin | mutant DELTAS183, Co+ assay | Limosilactobacillus reuteri | |
2.5.1.17 | 0.00011 | - |
cob(I)alamin | mutant F187A, Co+ assay | Limosilactobacillus reuteri | |
2.5.1.17 | 0.00013 | - |
cob(I)alamin | wild-type, Co+ assay | Limosilactobacillus reuteri | |
2.5.1.17 | 0.00019 | - |
cob(I)alamin | mutant F163A, Co+ assay | Limosilactobacillus reuteri | |
2.5.1.17 | 0.00055 | - |
cob(I)alamin | mutant F112Y, Co+ assay | Limosilactobacillus reuteri | |
2.5.1.17 | 0.00077 | - |
cob(I)alamin | mutant F112W, Co+ assay | Limosilactobacillus reuteri | |
2.5.1.17 | 0.0012 | - |
ATP | mutant F112Y, Co+ assay | Limosilactobacillus reuteri | |
2.5.1.17 | 0.0012 | - |
ATP | mutant F187A, Co+ assay | Limosilactobacillus reuteri | |
2.5.1.17 | 0.0019 | - |
cob(I)alamin | mutant F112A, Co+ assay | Limosilactobacillus reuteri | |
2.5.1.17 | 0.002 | - |
ATP | mutant F112W, Co+ assay | Limosilactobacillus reuteri | |
2.5.1.17 | 0.0022 | - |
ATP | wild-type, Co+ assay | Limosilactobacillus reuteri | |
2.5.1.17 | 0.0026 | - |
ATP | mutant F112H, Co+ assay | Limosilactobacillus reuteri | |
2.5.1.17 | 0.0028 | - |
cob(I)alamin | mutant F112H, Co+ assay | Limosilactobacillus reuteri | |
2.5.1.17 | 0.0031 | - |
ATP | mutant DELTAS183, Co+ assay | Limosilactobacillus reuteri | |
2.5.1.17 | 0.005 | - |
ATP | wild-type, Co2+ assay | Limosilactobacillus reuteri | |
2.5.1.17 | 0.0078 | - |
cob(II)alamin | wild-type, Co2+ assay | Limosilactobacillus reuteri | |
2.5.1.17 | 0.0079 | - |
ATP | mutant F187A, Co2+ assay | Limosilactobacillus reuteri | |
2.5.1.17 | 0.0099 | - |
ATP | mutant DELTAS183, Co2+ assay | Limosilactobacillus reuteri | |
2.5.1.17 | 0.0104 | - |
ATP | mutant F112H, Co2+ assay | Limosilactobacillus reuteri | |
2.5.1.17 | 0.015 | - |
cob(II)alamin | mutant F187A, Co2+ assay | Limosilactobacillus reuteri | |
2.5.1.17 | 0.0159 | - |
ATP | mutant F163A, Co+ assay | Limosilactobacillus reuteri | |
2.5.1.17 | 0.016 | - |
cob(II)alamin | mutant DELTAS183, Co2+ assay | Limosilactobacillus reuteri | |
2.5.1.17 | 0.028 | - |
cob(II)alamin | mutant F112W, Co2+ assay | Limosilactobacillus reuteri | |
2.5.1.17 | 0.034 | - |
cob(II)alamin | mutant F112Y, Co2+ assay | Limosilactobacillus reuteri | |
2.5.1.17 | 0.0346 | - |
ATP | mutant F112A, Co+ assay | Limosilactobacillus reuteri | |
2.5.1.17 | 0.053 | - |
cob(II)alamin | mutant F112H, Co2+ assay | Limosilactobacillus reuteri | |
2.5.1.17 | 0.073 | - |
ATP | mutant F112W, Co2+ assay | Limosilactobacillus reuteri | |
2.5.1.17 | 0.094 | - |
ATP | mutant F112Y, Co2+ assay | Limosilactobacillus reuteri | |
2.5.1.17 | 0.0961 | - |
ATP | mutant F163A, Co2+ assay | Limosilactobacillus reuteri | |
2.5.1.17 | 0.134 | - |
cob(II)alamin | mutant F163A, Co2+ assay | Limosilactobacillus reuteri |
EC Number | Organism | UniProt | Comment | Textmining |
---|---|---|---|---|
2.5.1.17 | Limosilactobacillus reuteri | - |
- |
- |
EC Number | Purification (Comment) | Organism |
---|---|---|
2.5.1.17 | using Ni-NTA chromatography | Limosilactobacillus reuteri |
EC Number | Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
---|---|---|---|---|---|---|---|
2.5.1.17 | cob(I)alamin + ATP | in the co+ assay the cobalt ion of cobalamin is chemically reduced in solution to cob(I)alamin by Ti(III)citrate, allowing the cob(I)alamin adenosylation reaction to be measured directly. The Co+ assay is performed under anoxic conditions | Limosilactobacillus reuteri | adenosylcobalamin + ? | - |
? | |
2.5.1.17 | cob(II)alamin + ATP | in the Co2+ assay the NADPH-dependent flavodoxin protein reductase/flavodoxin system is used to reduce Co2+ to Co+. In this Co2+ assay, the PduO enzyme must bind cob(II)alamin and facilitate the generation of cob(I)alamin in its active site | Limosilactobacillus reuteri | adenosylcobalamin + ? | - |
? |
EC Number | Synonyms | Comment | Organism |
---|---|---|---|
2.5.1.17 | LrPduO | - |
Limosilactobacillus reuteri |
EC Number | Temperature Optimum [°C] | Temperature Optimum Maximum [°C] | Comment | Organism |
---|---|---|---|---|
2.5.1.17 | 37 | - |
assay at | Limosilactobacillus reuteri |
EC Number | Turnover Number Minimum [1/s] | Turnover Number Maximum [1/s] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|---|
2.5.1.17 | 0.00038 | - |
ATP | mutant F112H, Co+ assay | Limosilactobacillus reuteri | |
2.5.1.17 | 0.00042 | - |
cob(I)alamin | mutant F112H, Co+ assay | Limosilactobacillus reuteri | |
2.5.1.17 | 0.0005 | - |
ATP | mutant F112H, Co2+ assay | Limosilactobacillus reuteri | |
2.5.1.17 | 0.00067 | - |
cob(II)alamin | mutant F112H, Co2+ assay | Limosilactobacillus reuteri | |
2.5.1.17 | 0.0016 | - |
ATP | mutant F112Y, Co+ assay | Limosilactobacillus reuteri | |
2.5.1.17 | 0.0017 | - |
cob(I)alamin | mutant F112Y, Co+ assay | Limosilactobacillus reuteri | |
2.5.1.17 | 0.0018 | - |
ATP | mutant F112W, Co+ assay | Limosilactobacillus reuteri | |
2.5.1.17 | 0.002 | - |
cob(I)alamin | mutant F112W, Co+ assay | Limosilactobacillus reuteri | |
2.5.1.17 | 0.0023 | - |
ATP | mutant F112Y, Co2+ assay | Limosilactobacillus reuteri | |
2.5.1.17 | 0.0027 | - |
cob(II)alamin | mutant F112Y, Co2+ assay | Limosilactobacillus reuteri | |
2.5.1.17 | 0.0028 | - |
ATP | mutant F112W, Co2+ assay | Limosilactobacillus reuteri | |
2.5.1.17 | 0.0032 | - |
cob(II)alamin | mutant F112W, Co2+ assay | Limosilactobacillus reuteri | |
2.5.1.17 | 0.0056 | - |
ATP | mutant F112A, Co+ assay | Limosilactobacillus reuteri | |
2.5.1.17 | 0.0066 | - |
cob(I)alamin | mutant F112A, Co+ assay | Limosilactobacillus reuteri | |
2.5.1.17 | 0.014 | - |
ATP | mutant F187A, Co2+ assay | Limosilactobacillus reuteri | |
2.5.1.17 | 0.015 | - |
ATP | mutant F163A, Co+ assay | Limosilactobacillus reuteri | |
2.5.1.17 | 0.015 | - |
cob(I)alamin | mutant F163A, Co+ assay | Limosilactobacillus reuteri | |
2.5.1.17 | 0.015 | - |
ATP | mutant F163A, Co2+ assay | Limosilactobacillus reuteri | |
2.5.1.17 | 0.015 | - |
cob(II)alamin | mutant F187A, Co2+ assay | Limosilactobacillus reuteri | |
2.5.1.17 | 0.017 | - |
ATP | mutant DELTAS183, Co2+ assay | Limosilactobacillus reuteri | |
2.5.1.17 | 0.018 | - |
cob(II)alamin | mutant DELTAS183, Co2+ assay | Limosilactobacillus reuteri | |
2.5.1.17 | 0.018 | - |
cob(I)alamin | mutant F187A, Co+ assay | Limosilactobacillus reuteri | |
2.5.1.17 | 0.02 | - |
cob(I)alamin | mutant DELTAS183, Co+ assay | Limosilactobacillus reuteri | |
2.5.1.17 | 0.021 | - |
ATP | mutant F187A, Co+ assay | Limosilactobacillus reuteri | |
2.5.1.17 | 0.023 | - |
ATP | mutant DELTAS183, Co+ assay | Limosilactobacillus reuteri | |
2.5.1.17 | 0.024 | - |
cob(I)alamin | wild-type, Co+ assay | Limosilactobacillus reuteri | |
2.5.1.17 | 0.026 | - |
ATP | wild-type, Co+ assay | Limosilactobacillus reuteri | |
2.5.1.17 | 0.027 | - |
cob(II)alamin | mutant F163A, Co2+ assay | Limosilactobacillus reuteri | |
2.5.1.17 | 0.029 | - |
ATP | wild-type, Co2+ assay | Limosilactobacillus reuteri | |
2.5.1.17 | 0.038 | - |
cob(II)alamin | wild-type, Co2+ assay | Limosilactobacillus reuteri |
EC Number | pH Optimum Minimum | pH Optimum Maximum | Comment | Organism |
---|---|---|---|---|
2.5.1.17 | 8 | - |
assay at | Limosilactobacillus reuteri |
EC Number | kcat/KM Value [1/mMs-1] | kcat/KM Value Maximum [1/mMs-1] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|---|
2.5.1.17 | 0.013 | - |
cob(II)alamin | mutant F112H, Co2+ assay | Limosilactobacillus reuteri | |
2.5.1.17 | 0.057 | - |
ATP | mutant F112H, Co2+ assay | Limosilactobacillus reuteri | |
2.5.1.17 | 0.083 | - |
cob(II)alamin | mutant F112Y, Co2+ assay | Limosilactobacillus reuteri | |
2.5.1.17 | 0.11 | - |
cob(II)alamin | mutant F112W, Co2+ assay | Limosilactobacillus reuteri | |
2.5.1.17 | 0.14 | - |
ATP | mutant F112H, Co+ assay | Limosilactobacillus reuteri | |
2.5.1.17 | 0.16 | - |
ATP | mutant F163A, Co2+ assay | Limosilactobacillus reuteri | |
2.5.1.17 | 0.17 | - |
ATP | mutant F112A, Co+ assay | Limosilactobacillus reuteri | |
2.5.1.17 | 0.2 | - |
cob(II)alamin | mutant F163A, Co2+ assay | Limosilactobacillus reuteri | |
2.5.1.17 | 0.25 | - |
ATP | mutant F112Y, Co2+ assay | Limosilactobacillus reuteri | |
2.5.1.17 | 0.39 | - |
ATP | mutant F112W, Co2+ assay | Limosilactobacillus reuteri | |
2.5.1.17 | 0.93 | - |
cob(II)alamin | mutant F187A, Co2+ assay | Limosilactobacillus reuteri | |
2.5.1.17 | 0.98 | - |
ATP | mutant F112W, Co+ assay | Limosilactobacillus reuteri | |
2.5.1.17 | 1.1 | - |
cob(II)alamin | mutant DELTAS183, Co2+ assay | Limosilactobacillus reuteri | |
2.5.1.17 | 1.4 | - |
ATP | mutant F112Y, Co+ assay | Limosilactobacillus reuteri | |
2.5.1.17 | 1.7 | - |
ATP | mutant DELTAS183, Co2+ assay | Limosilactobacillus reuteri | |
2.5.1.17 | 1.8 | - |
ATP | mutant F163A, Co+ assay | Limosilactobacillus reuteri | |
2.5.1.17 | 1.8 | - |
ATP | mutant F187A, Co2+ assay | Limosilactobacillus reuteri | |
2.5.1.17 | 2.6 | - |
cob(I)alamin | mutant F112W, Co+ assay | Limosilactobacillus reuteri | |
2.5.1.17 | 3.1 | - |
cob(I)alamin | mutant F112Y, Co+ assay | Limosilactobacillus reuteri | |
2.5.1.17 | 3.6 | - |
cob(I)alamin | mutant F112A, Co+ assay | Limosilactobacillus reuteri | |
2.5.1.17 | 4.8 | - |
cob(II)alamin | wild-type, Co2+ assay | Limosilactobacillus reuteri | |
2.5.1.17 | 5.5 | - |
ATP | wild-type, Co2+ assay | Limosilactobacillus reuteri | |
2.5.1.17 | 7.5 | - |
ATP | mutant DELTAS183, Co+ assay | Limosilactobacillus reuteri | |
2.5.1.17 | 12 | - |
ATP | wild-type, Co+ assay | Limosilactobacillus reuteri | |
2.5.1.17 | 15 | - |
cob(I)alamin | mutant F112H, Co+ assay | Limosilactobacillus reuteri | |
2.5.1.17 | 17 | - |
ATP | mutant F187A, Co+ assay | Limosilactobacillus reuteri | |
2.5.1.17 | 79 | - |
cob(I)alamin | mutant F163A, Co+ assay | Limosilactobacillus reuteri | |
2.5.1.17 | 160 | - |
cob(I)alamin | mutant F187A, Co+ assay | Limosilactobacillus reuteri | |
2.5.1.17 | 180 | - |
cob(I)alamin | wild-type, Co+ assay | Limosilactobacillus reuteri | |
2.5.1.17 | 395 | - |
cob(I)alamin | mutant DELTAS183, Co+ assay | Limosilactobacillus reuteri |