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Literature summary for 1.17.1.10 extracted from

  • Hatrongjit, R.; Packdibamrung, K.
    A novel NADP+-dependent formate dehydrogenase from Burkholderia stabilis 15516: Screening, purification and characterization (2010), Enzyme Microb. Technol., 46, 557-561.
No PubMed abstract available
Search for more literature for 1.17.1.10

Cloned(Commentary)

Cloned (Comment) Organism
expressed in Escherichia coli BL21(DE3) cells Burkholderia stabilis

Protein Variants

Protein Variants Comment Organism
Q223E the mutated enzyme shows an 18fold increment in catalytic efficiency with NAD+ with a concomitant 185fold reduction in the efficiency with NADP+, drastically shifting the cofactor preference from NADP+ to NAD+ Burkholderia stabilis

Inhibitors

Inhibitors Comment Organism Structure
5,5'-dithio-bis(2-nitrobenzoic acid) complete inhibition at 1 mM Burkholderia stabilis
AgNO3 complete inhibition at 1 mM Burkholderia stabilis
CdCl2 slight inhibition at 1 mM Burkholderia stabilis
CoCl2 slight inhibition at 1 mM Burkholderia stabilis
CuSO4 slight inhibition at 1 mM Burkholderia stabilis
FeCl2 complete inhibition at 1 mM Burkholderia stabilis
FeSO4 slight inhibition at 1 mM Burkholderia stabilis
HgCl2 complete inhibition at 1 mM Burkholderia stabilis
hydroxylamine slight inhibition at 10 mM Burkholderia stabilis
additional information ZnCl2 (1 mM), 10 mM of Mg2SO4, dithiothreitol, mercaptoethanol and phenylmethylsulfonyl fluoride do not affect enzyme activity Burkholderia stabilis
NaNO3 slight inhibition at 1 mM Burkholderia stabilis
Sodium azide complete inhibition at 1 mM Burkholderia stabilis

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
0.06
-
 NAD+ mutant enzyme Q223E, in 1 M potassium phosphate buffer (pH 7.0), at 30°C Burkholderia stabilis
0.16
-
 NADP+ wild type enzyme, in 1 M potassium phosphate buffer (pH 7.0), at 30°C Burkholderia stabilis
0.83
-
 NADP+ mutant enzyme Q223E, in 1 M potassium phosphate buffer (pH 7.0), at 30°C Burkholderia stabilis
1.43
-
 NAD+ wild type enzyme, in 1 M potassium phosphate buffer (pH 7.0), at 30°C Burkholderia stabilis
55.5
-
 formate wild type enzyme, in 1 M potassium phosphate buffer (pH 7.0), at 30°C Burkholderia stabilis

Molecular Weight [Da]

Molecular Weight [Da] Molecular Weight Maximum [Da] Comment Organism
42000
-
 8 * 42000, SDS-PAGE Burkholderia stabilis
340000
-
 gel filtration Burkholderia stabilis

Organism

Organism UniProt Comment Textmining
Burkholderia stabilis B5A8W5
-
-
Burkholderia stabilis 15516 B5A8W5
-
-

Purification (Commentary)

Purification (Comment) Organism
DEAE-Toyopearl column chromatography and butyl-Toyopearl 650M column chromatography Burkholderia stabilis

Specific Activity [micromol/min/mg]

Specific Activity Minimum [µmol/min/mg] Specific Activity Maximum [µmol/min/mg] Comment Organism
1.4
-
 crude enzyme, in 1 M potassium phosphate buffer (pH 7.0), at 30°C Burkholderia stabilis
5.3
-
 after 3.8fold purification, in 1 M potassium phosphate buffer (pH 7.0), at 30°C Burkholderia stabilis

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
formate + NAD+ the activity to NAD+ is 38% of that to NADP+ Burkholderia stabilis CO2 + NADH + H+
-
 ?
formate + NAD+ the activity to NAD+ is 38% of that to NADP+ Burkholderia stabilis 15516 CO2 + NADH + H+
-
 ?
formate + NADP+ the enzyme acts only on formate as the substrate Burkholderia stabilis CO2 + NADPH + H+
-
 ?
formate + NADP+ the enzyme acts only on formate as the substrate Burkholderia stabilis 15516 CO2 + NADPH + H+
-
 ?
additional information methanol, formaldehyde, acetic acid, ethanol, propanol and glycerol are inert. When artificial electron acceptors are used, ferricyanide, nitro blue tetrazolium, phenazine methosulfate, riboflavin, ATP and FAD are not reduced in the presence of formate Burkholderia stabilis ?
-
 ?
additional information methanol, formaldehyde, acetic acid, ethanol, propanol and glycerol are inert. When artificial electron acceptors are used, ferricyanide, nitro blue tetrazolium, phenazine methosulfate, riboflavin, ATP and FAD are not reduced in the presence of formate Burkholderia stabilis 15516 ?
-
 ?

Subunits

Subunits Comment Organism
homooctamer 8 * 42000, SDS-PAGE Burkholderia stabilis

Synonyms

Synonyms Comment Organism
FDH
-
 Burkholderia stabilis
NADP+-dependent formate dehydrogenase
-
 Burkholderia stabilis

Temperature Optimum [°C]

Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
60
-
-
 Burkholderia stabilis

Temperature Stability [°C]

Temperature Stability Minimum [°C] Temperature Stability Maximum [°C] Comment Organism
55 60 no loss of activity is observed at 55°C for 10 h, but about half of the enzyme activity is lost after incubation for 36 h. The enzyme can maintain a 50% activity for at least 5 h when it is incubated at 60°C Burkholderia stabilis

Turnover Number [1/s]

Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
0.13
-
 NADP+ mutant enzyme Q223E, in 1 M potassium phosphate buffer (pH 7.0), at 30°C Burkholderia stabilis
1.25
-
 NAD+ mutant enzyme Q223E, in 1 M potassium phosphate buffer (pH 7.0), at 30°C Burkholderia stabilis
1.66
-
 NAD+ wild type enzyme, in 1 M potassium phosphate buffer (pH 7.0), at 30°C Burkholderia stabilis
4.75
-
 NADP+ wild type enzyme, in 1 M potassium phosphate buffer (pH 7.0), at 30°C Burkholderia stabilis

pH Optimum

pH Optimum Minimum pH Optimum Maximum Comment Organism
6 7.5
-
 Burkholderia stabilis

pH Stability

pH Stability pH Stability Maximum Comment Organism
5 12 more than 94% of the initial activity is retained at pH 5.0-12.0 after incubation for 30 min at 30°C Burkholderia stabilis

Cofactor

Cofactor Comment Organism Structure
NADP+ the enzyme accepts NADP+ as the natural coenzyme with a much higher specificity over NAD+, the activity to NAD+ is 38% of that to NADP+ Burkholderia stabilis

kcat/KM [mM/s]

kcat/KM Value [1/mMs-1] kcat/KM Value Maximum [1/mMs-1] Substrate Comment Organism Structure
0.16
-
 NADP+ mutant enzyme Q223E, in 1 M potassium phosphate buffer (pH 7.0), at 30°C Burkholderia stabilis
1.16
-
 NAD+ wild type enzyme, in 1 M potassium phosphate buffer (pH 7.0), at 30°C Burkholderia stabilis
20
-
 NAD+ mutant enzyme Q223E, in 1 M potassium phosphate buffer (pH 7.0), at 30°C Burkholderia stabilis
30
-
 NADP+ wild type enzyme, in 1 M potassium phosphate buffer (pH 7.0), at 30°C Burkholderia stabilis