Benzoylformate dehydrogenase from Pseudomonas putida

Enzyme Description

Species
Pseudomonas putida β†—
Extremophile
No
EC Number
4.1.1.7 β†—

Sequence

Length: 528 amino acids
MASVHGTTYELLRRQGIDTVFGNPGSNELPFLKDFPEDFRYILALQEACVVGIADGYAQASRKPAFINLHSAAGTGNAMGALSNAWNSHSPLIVTAGQQTRAMIGVEALLTNVDAANLPRPLVKWSYEPASAAEVPHAMSRAIHMASMAPQGPVYLSVPYDDWDKDADPQSHHLFDRHVSSSVRLNDQDLDILVKALNSASNPAIVLGPDVDAANANADCVMLAERLKAPVWVAPSAPRCPFPTRHPCFRGLMPAGIAAISQLLEGHDVVLVIGAPVFRYHQYDPGQYLKPGTRLISVTCDPLEAARAPMGDAIVADIGAMASALANLVEESSRQLPTAAPEPAKVDQDAGRLHPETVFDTLNDMAPENAIYLNESTSTTAQMWQRLNMRNPGSYYFCAAGGLGFALPAAIGVQLAEPERQVIAVIGDGSANYSISALWTAAQYNIPTIFVIMNNGTYGALRWFAGVLEAENVPGLDVPGIDFRALAKGYGVQALKADNLEQLKGSLQEALSAKGPVLIEVSTVSPVK
B. Lingen et al. (2002) β€” Improving the carboligase activity of benzoylformate decarboxylase from Pseudomonas putida by a combination of directed evolution and site-directed mutagenesis
Protein Engineering, Design and Selection  Β· doi:10.1093/protein/15.7.585 β†—  Β· Activity - Classical
22 measurements ▼
Database ID
UniProt: P20906 β†—
Sequence Annotation
Inferred - from protein name
Protein Source
Recombinant, host bacterium Escherichia coli SG13009

Experimental Data (22 measurements)

22 measurements β€” page 1 of 2
Mutation Property Assay Solvent Solvent Volume Aqueous ReferenceWT Reference Measured Value Units Extraction method Data type Solution pH Temperature Substrate(s) Product(s) Cofactor(s) Shaking Comments
L476M Activity - Classical Activity (as a percentage of the WT) measured by HPLC in the presence of organic solvent Dimethyl Sulfoxide (DMSO) 20 % v/v 382 100 423 % Direct Continuous 50 mM potassium phosphate buffer, 2.5 mM MgSO4 7 30Β°C 500 mM Acetaldehyde, 40 mM Benzaldehyde 1-Hydroxy-1-phenyl-2-propanone 0.5 mM ThDP β€” Classical aqueous control (in %) | Control compared to WT activity
L476G Activity - Classical Activity (as a percentage of the WT) measured by HPLC in the presence of organic solvent Dimethyl Sulfoxide (DMSO) 20 % v/v 237 100 427 % Direct Continuous 50 mM potassium phosphate buffer, 2.5 mM MgSO4 7 30Β°C 500 mM Acetaldehyde, 40 mM Benzaldehyde 1-Hydroxy-1-phenyl-2-propanone 0.5 mM ThDP β€” Classical aqueous control (in %) | Control compared to WT activity
L476G Activity - Classical Activity (as a percentage of the WT) measured by HPLC in the presence of organic solvent Ethanol 1.5 Molar 237 100 469 % Direct Continuous 50 mM potassium phosphate buffer, 2.5 mM MgSO4 7 30Β°C 500 mM Acetaldehyde, 40 mM Benzaldehyde 1-Hydroxy-1-phenyl-2-propanone 0.5 mM ThDP β€” Classical aqueous control (in %) | Control compared to WT activity
L476H Activity - Classical Activity (as a percentage of the WT) measured by HPLC in the presence of organic solvent Dimethyl Sulfoxide (DMSO) 20 % v/v 203 100 319 % Direct Continuous 50 mM potassium phosphate buffer, 2.5 mM MgSO4 7 30Β°C 500 mM Acetaldehyde, 40 mM Benzaldehyde 1-Hydroxy-1-phenyl-2-propanone 0.5 mM ThDP β€” Classical aqueous control (in %) | Control compared to WT activity
L476H Activity - Classical Activity (as a percentage of the WT) measured by HPLC in the presence of organic solvent Ethanol 1.5 Molar 203 100 362 % Direct Continuous 50 mM potassium phosphate buffer, 2.5 mM MgSO4 7 30Β°C 500 mM Acetaldehyde, 40 mM Benzaldehyde 1-Hydroxy-1-phenyl-2-propanone 0.5 mM ThDP β€” Classical aqueous control (in %) | Control compared to WT activity
L476A Activity - Classical Activity (as a percentage of the WT) measured by HPLC in the presence of organic solvent Dimethyl Sulfoxide (DMSO) 20 % v/v 244 100 411 % Direct Continuous 50 mM potassium phosphate buffer, 2.5 mM MgSO4 7 30Β°C 500 mM Acetaldehyde, 40 mM Benzaldehyde 1-Hydroxy-1-phenyl-2-propanone 0.5 mM ThDP β€” Classical aqueous control (in %) | Control compared to WT activity
L476A Activity - Classical Activity (as a percentage of the WT) measured by HPLC in the presence of organic solvent Ethanol 1.5 Molar 244 100 558 % Direct Continuous 50 mM potassium phosphate buffer, 2.5 mM MgSO4 7 30Β°C 500 mM Acetaldehyde, 40 mM Benzaldehyde 1-Hydroxy-1-phenyl-2-propanone 0.5 mM ThDP β€” Classical aqueous control (in %) | Control compared to WT activity
L476K Activity - Classical Activity (as a percentage of the WT) measured by HPLC in the presence of organic solvent Dimethyl Sulfoxide (DMSO) 20 % v/v 292 100 500 % Direct Continuous 50 mM potassium phosphate buffer, 2.5 mM MgSO4 7 30Β°C 500 mM Acetaldehyde, 40 mM Benzaldehyde 1-Hydroxy-1-phenyl-2-propanone 0.5 mM ThDP β€” Classical aqueous control (in %) | Control compared to WT activity
L476K Activity - Classical Activity (as a percentage of the WT) measured by HPLC in the presence of organic solvent Ethanol 1.5 Molar 292 100 592 % Direct Continuous 50 mM potassium phosphate buffer, 2.5 mM MgSO4 7 30Β°C 500 mM Acetaldehyde, 40 mM Benzaldehyde 1-Hydroxy-1-phenyl-2-propanone 0.5 mM ThDP β€” Classical aqueous control (in %) | Control compared to WT activity
L476T Activity - Classical Activity (as a percentage of the WT) measured by HPLC in the presence of organic solvent Dimethyl Sulfoxide (DMSO) 20 % v/v 328 100 398 % Direct Continuous 50 mM potassium phosphate buffer, 2.5 mM MgSO4 7 30Β°C 500 mM Acetaldehyde, 40 mM Benzaldehyde 1-Hydroxy-1-phenyl-2-propanone 0.5 mM ThDP β€” Classical aqueous control (in %) | Control compared to WT activity
L476T Activity - Classical Activity (as a percentage of the WT) measured by HPLC in the presence of organic solvent Ethanol 1.5 Molar 328 100 500 % Direct Continuous 50 mM potassium phosphate buffer, 2.5 mM MgSO4 7 30Β°C 500 mM Acetaldehyde, 40 mM Benzaldehyde 1-Hydroxy-1-phenyl-2-propanone 0.5 mM ThDP β€” Classical aqueous control (in %) | Control compared to WT activity
L476Q Activity - Classical Activity (as a percentage of the WT) measured by HPLC in the presence of organic solvent Ethanol 1.5 Molar 395 100 727 % Direct Continuous 50 mM potassium phosphate buffer, 2.5 mM MgSO4 7 30Β°C 500 mM Acetaldehyde, 40 mM Benzaldehyde 1-Hydroxy-1-phenyl-2-propanone 0.5 mM ThDP β€” Classical aqueous control (in %) | Control compared to WT activity
L476M Activity - Classical Activity (as a percentage of the WT) measured by HPLC in the presence of organic solvent Ethanol 1.5 Molar 382 100 562 % Direct Continuous 50 mM potassium phosphate buffer, 2.5 mM MgSO4 7 30Β°C 500 mM Acetaldehyde, 40 mM Benzaldehyde 1-Hydroxy-1-phenyl-2-propanone 0.5 mM ThDP β€” Classical aqueous control (in %) | Control compared to WT activity
L476C Activity - Classical Activity (as a percentage of the WT) measured by HPLC in the presence of organic solvent Dimethyl Sulfoxide (DMSO) 20 % v/v 270 100 431 % Direct Continuous 50 mM potassium phosphate buffer, 2.5 mM MgSO4 7 30Β°C 500 mM Acetaldehyde, 40 mM Benzaldehyde 1-Hydroxy-1-phenyl-2-propanone 0.5 mM ThDP β€” Classical aqueous control (in %) | Control compared to WT activity
L476C Activity - Classical Activity (as a percentage of the WT) measured by HPLC in the presence of organic solvent Ethanol 1.5 Molar 270 100 515 % Direct Continuous 50 mM potassium phosphate buffer, 2.5 mM MgSO4 7 30Β°C 500 mM Acetaldehyde, 40 mM Benzaldehyde 1-Hydroxy-1-phenyl-2-propanone 0.5 mM ThDP β€” Classical aqueous control (in %) | Control compared to WT activity
L476S Activity - Classical Activity (as a percentage of the WT) measured by HPLC in the presence of organic solvent Dimethyl Sulfoxide (DMSO) 20 % v/v 290 100 350 % Direct Continuous 50 mM potassium phosphate buffer, 2.5 mM MgSO4 7 30Β°C 500 mM Acetaldehyde, 40 mM Benzaldehyde 1-Hydroxy-1-phenyl-2-propanone 0.5 mM ThDP β€” Classical aqueous control (in %) | Control compared to WT activity
L476S Activity - Classical Activity (as a percentage of the WT) measured by HPLC in the presence of organic solvent Ethanol 1.5 Molar 290 100 420 % Direct Continuous 50 mM potassium phosphate buffer, 2.5 mM MgSO4 7 30Β°C 500 mM Acetaldehyde, 40 mM Benzaldehyde 1-Hydroxy-1-phenyl-2-propanone 0.5 mM ThDP β€” Classical aqueous control (in %) | Control compared to WT activity
S181T/L476P Activity - Classical Activity (as a percentage of the WT) measured by HPLC in the presence of organic solvent Dimethyl Sulfoxide (DMSO) 20 % v/v 66 100 175 % Direct Continuous 50 mM potassium phosphate buffer, 2.5 mM MgSO4 7 30Β°C 500 mM Acetaldehyde, 40 mM Benzaldehyde 1-Hydroxy-1-phenyl-2-propanone 0.5 mM ThDP β€” Classical aqueous control (in %) | Control compared to WT activity
S181T/L476P Activity - Classical Activity (as a percentage of the WT) measured by HPLC in the presence of organic solvent Ethanol 1.5 Molar 66 100 142 % Direct Continuous 50 mM potassium phosphate buffer, 2.5 mM MgSO4 7 30Β°C 500 mM Acetaldehyde, 40 mM Benzaldehyde 1-Hydroxy-1-phenyl-2-propanone 0.5 mM ThDP β€” Classical aqueous control (in %) | Control compared to WT activity
L476Q/S525G Activity - Classical Activity (as a percentage of the WT) measured by HPLC in the presence of organic solvent Dimethyl Sulfoxide (DMSO) 20 % v/v 245 100 325 % Direct Continuous 50 mM potassium phosphate buffer, 2.5 mM MgSO4 7 30Β°C 500 mM Acetaldehyde, 40 mM Benzaldehyde 1-Hydroxy-1-phenyl-2-propanone 0.5 mM ThDP β€” Classical aqueous control (in %) | Control compared to WT activity
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Mutations in this dataset (11)

S181T/L476P L476A L476C L476G L476H L476K L476M L476Q L476Q/S525G L476S L476T

Visualization : Activity β€” Classical

One bar per measurement. Colour = solvent, shade = solvent volume. β€” β€” β€” Reference value. Hover for details.

Mutation Effect

Mutation impact on enzyme stability and function in the presence of organic solvent: comparison of wild-type and mutant values in identical conditions.

Structure

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