Cloned (Comment) | Organism |
---|---|
expression of mutant enzyme L476Q and M365L/L461S in Escherichia coli | Pseudomonas putida |
Protein Variants | Comment | Organism |
---|---|---|
L476Q | mutant enzyme selectively catalyzes the formation of enantiopure (S)-2-hydroxy-1-(2-methylphenyl)propan-1-one with excellent yield, a reaction which is only poorly catalyzed by the wild-type enzyme. Vmax and Km-value for decraboxylation of benzoylformate are not effected | Pseudomonas putida |
M365L/L461S | mutant enzyme selectively catalyzes the formation of enantiopure (S)-2-hydroxy-1-(2-methylphenyl)propan-1-one with excellent yield, a reaction which is only poorly catalyzed by the wild-type enzyme. Mutant enzyme retains only 9% of the wild-type BFD carboligase activity. Decrease in Vmax value as well as an increase in the Km-value for decarboxylation of benzoylformate by mutant enzyme compared to that of wild-type enzyme | Pseudomonas putida |
Inhibitors | Comment | Organism | Structure |
---|---|---|---|
benzoylformate | substrate inhibition | Pseudomonas putida |
KM Value [mM] | KM Value Maximum [mM] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
0.54 | - |
benzoylformate | wild-type enzyme | Pseudomonas putida | |
0.58 | - |
benzoylformate | mutant enzyme L476Q | Pseudomonas putida |
Organism | UniProt | Comment | Textmining |
---|---|---|---|
Pseudomonas putida | P20906 | - |
- |
Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
---|---|---|---|---|---|---|
benzoylformate | - |
Pseudomonas putida | benzaldehyde + CO2 | - |
? | |
m-bromo-benzoylformate | wild-type enzyme: 68% conversion with 96% enantiomeric excess of the (S)-2-hydroxy ketone, mutant enzyme L476Q: 97% conversion with more than 98% enantiomeric excess of the (S)-2-hydroxy ketone, mutant enzyme M365L-L461S: 100% conversion with 98.5% enantiomeric excess of the (S)-2-hydroxy ketone | Pseudomonas putida | m-bromo-benzaldehyde + CO2 | - |
? | |
m-chloro-benzoylformate | wild-type enzyme: 94% conversion with 94% enantiomeric excess of the (S)-2-hydroxy ketone, mutant enzyme L476Q: 100% conversion with more than 97% enantiomeric excess of the (S)-2-hydroxy ketone, mutant enzyme M365L-L461S: 100% conversion with 98% enantiomeric excess of the (S)-2-hydroxy ketone | Pseudomonas putida | m-chloro-benzaldehyde + CO2 | - |
? | |
m-fluoro-benzoylformate | wild-type enzyme: 100% conversion with 87% enantiomeric excess of the (S)-2-hydroxy ketone, mutant enzyme L476Q: 100% conversion with more than 99% enantiomeric excess of the (S)-2-hydroxy ketone, mutant enzyme M365L-L461S: 94% conversion with more than 99% enantiomeric excess of the (S)-2-hydroxy ketone | Pseudomonas putida | m-fluoro-benzaldehyde + CO2 | - |
? | |
m-methoxy-benzoylformate | wild-type enzyme: 94% conversion with 96% enantiomeric excess of the (S)-2-hydroxy ketone, mutant enzyme L476Q: 100% conversion with more than 99% enantiomeric excess of the (S)-2-hydroxy ketone, mutant enzyme M365L-L461S: 100% conversion with 99% enantiomeric excess of the (S)-2-hydroxy ketone | Pseudomonas putida | m-methoxy-benzaldehyde + CO2 | - |
? | |
m-methyl-benzoylformate | wild-type enzyme: 99% conversion with 97% enantiomeric excess of the (S)-2-hydroxy ketone, mutant enzyme L476Q: 100% conversion with more than 99% enantiomeric excess of the (S)-2-hydroxy ketone, mutant enzyme M365L-L461S: 100% conversion with 99% enantiomeric excess of the (S)-2-hydroxy ketone | Pseudomonas putida | m-methyl-benzaldehyde + CO2 | - |
? | |
o-bromo-benzoylformate | wild-type enzyme: no activity, mutant enzyme L476Q: 98% conversion with more than 99% enantiomeric excess of the (S)-2-hydroxy ketone, mutant enzyme M365L-L461S: 90% conversion with 99% enantiomeric excess of the (S)-2-hydroxy ketone | Pseudomonas putida | o-bromo-benzaldehyde + CO2 | - |
? | |
o-chloro-benzoylformate | wild-type enzyme: no activity, mutant enzyme L476Q: 100% conversion with more than 99% enantiomeric excess of the (S)-2-hydroxy ketone, mutant enzyme M365L-L461S: 85% conversion with more than 99% enantiomeric excess of the (S)-2-hydroxy ketone | Pseudomonas putida | o-chloro-benzaldehyde + CO2 | - |
? | |
o-fluoro-benzoylformate | wild-type enzyme: 91% conversion with 89% enantiomeric excess of the (S)-2-hydroxy ketone, mutant enzyme L476Q: 100% conversion with more than 99% enantiomeric excess of the (S)-2-hydroxy ketone, mutant enzyme M365L-L461S: 83% conversion with 98% enantiomeric excess of the (S)-2-hydroxy ketone | Pseudomonas putida | o-fluoro-benzaldehyde + CO2 | - |
? | |
o-methoxy-benzoylformate | wild-type enzyme: no conversion, mutant enzyme L476Q: 97% conversion with more than 99% enantiomeric excess of the (S)-2-hydroxy ketone, mutant enzyme M365L-L461S: 46% conversion with 99% enantiomeric excess of the (S)-2-hydroxy ketone | Pseudomonas putida | o-methoxy-benzaldehyde + CO2 | - |
? | |
o-methyl-benzoylformate | wild-type enzyme: 4% conversion, mutant enzyme L476Q: 100% conversion with more than 99% enantiomeric excess of the (S)-2-hydroxy ketone, mutant enzyme M365L-L461S: 83% conversion with 98% enantiomeric excess of the (S)-2-hydroxy ketone | Pseudomonas putida | o-methyl-benzaldehyde + CO2 | - |
? | |
o-methylbenzaldehyde + acetaldehyde | mutant enzyme L476Q and M365L/L461S selectively catalyzes the formation of enantiopure (S)-2-hydroxy-1-(2-methylphenyl)propan-1-one with excellent yield, a reaction which is only poorly catalyzed by the wild-type enzyme | Pseudomonas putida | (2S)-2-hydroxy-1-(2-methylphenyl)propan-1-one | - |
? | |
p-bromo-benzoylformate | wild-type enzyme: 42% conversion with 83% enantiomeric excess of the (S)-2-hydroxy ketone, mutant enzyme L476Q: 100% conversion with 96.5% enantiomeric excess of the (S)-2-hydroxy ketone, mutant enzyme M365L-L461S: 100% conversion with 96% enantiomeric excess of the (S)-2-hydroxy ketone | Pseudomonas putida | p-bromo-benzaldehyde + CO2 | - |
? | |
p-chloro-benzoylformate | wild-type enzyme: 85% conversion with 82% enantiomeric excess of the (S)-2-hydroxy ketone, mutant enzyme L476Q: 100% conversion with more than 96.5% enantiomeric excess of the (S)-2-hydroxy ketone, mutant enzyme M365L-L461S: 100% conversion with 95.5% enantiomeric excess of the (S)-2-hydroxy ketone | Pseudomonas putida | p-chloro-benzaldehyde + CO2 | - |
? | |
p-fluoro-benzoylformate | wild-type enzyme: 69% conversion with 87% enantiomeric excess of the (S)-2-hydroxy ketone, mutant enzyme L476Q: 100% conversion with 97% enantiomeric excess of the (S)-2-hydroxy ketone, mutant enzyme M365L-L461S: 87% conversion with 97% enantiomeric excess of the (S)-2-hydroxy ketone | Pseudomonas putida | p-fluoro-benzaldehyde + CO2 | - |
? | |
p-methoxy-benzoylformate | wild-type enzyme: 23% conversion with 92% enantiomeric excess of the (S)-2-hydroxy ketone, mutant enzyme L476Q: 100% conversion with more than 99% enantiomeric excess of the (S)-2-hydroxy ketone, mutant enzyme M365L-L461S: 67% conversion with 42% enantiomeric excess of the (S)-2-hydroxy ketone | Pseudomonas putida | p-methoxy-benzaldehyde + CO2 | - |
? | |
p-methyl-benzoylformate | wild-type enzyme: 65% conversion with 88% enantiomeric excess of the (S)-2-hydroxy ketone, mutant enzyme L476Q: 100% conversion with more than 98% enantiomeric excess of the (S)-2-hydroxy ketone, mutant enzyme M365L-L461S: 98% conversion with 98% enantiomeric excess of the (S)-2-hydroxy ketone | Pseudomonas putida | p-methyl-benzaldehyde + CO2 | - |
? |
Ki Value [mM] | Ki Value maximum [mM] | Inhibitor | Comment | Organism | Structure |
---|---|---|---|---|---|
100 | - |
benzoylformate | mutant enzyme L476Q | Pseudomonas putida | |
143 | - |
benzoylformate | wild-type enzyme | Pseudomonas putida |