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Literature summary for 1.20.1.1 extracted from

  • Fogle, E.J.; van der Donk, W.A.
    Pre-steady-state studies of phosphite dehydrogenase demonstrate that hydride transfer is fully rate limiting (2007), Biochemistry, 46, 13101-13108.
    View publication on PubMedView publication on EuropePMC

Application

Application Comment Organism
additional information has great potential for cofactor regeneration Pseudomonas stutzeri

Cloned(Commentary)

Cloned (Comment) Organism
His6-tagged PTDH overexpressed in Escherichia coli BL21(DE3) harboring a pET15b vector Pseudomonas stutzeri

Protein Variants

Protein Variants Comment Organism
D79A significant differences in its kinetic constants compared to the wild-type enzyme. 2600fold decrease in catalytic efficiency. Pre-steady-state rates are approximately the same as the steady-state rates Pseudomonas stutzeri
D79N has kinetic parameters more similar to those of wild-type Pseudomonas stutzeri
E266Q higher activity, steady-state and pre-steady-state rates are comparable Pseudomonas stutzeri
K76A pre-steady-state rates are approximately the same as the steady-state rates Pseudomonas stutzeri
additional information thermostable mutant 12X-PTDH with higher solubility than the wild-type. Thermostable mutant with dual cofactor specificity NADP-12X NAD shows pre-steady-state behavior very similar to that observed with 12X-PTDH and the wild-type enzyme. Pre-steady-state traces of thermostable mutant with dual cofactor specificity NADP-12X NADP shows curvature with NADP, particularly with protiated phosphite Pseudomonas stutzeri
R237K low activity, absence of a significant burst in the pre-steady-state Pseudomonas stutzeri

Inhibitors

Inhibitors Comment Organism Structure
additional information slow steps after hydride transfer do not significantly limit the rate of reaction for the wild-type enzyme, the active site mutants, or the thermostable mutant Pseudomonas stutzeri

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
0.001
-
phosphite mutant R237K Pseudomonas stutzeri
0.012
-
NAD+ mutant D79N Pseudomonas stutzeri
0.035
-
NAD+ wild-type Pseudomonas stutzeri
0.035
-
phosphite mutant D79N Pseudomonas stutzeri
0.055
-
phosphite wild-type Pseudomonas stutzeri
0.062
-
NAD+ mutant D79A Pseudomonas stutzeri
1
-
NAD+ mutant R237K Pseudomonas stutzeri
1.2
-
phosphite mutant D79A Pseudomonas stutzeri

Organism

Organism UniProt Comment Textmining
Pseudomonas stutzeri
-
-
-
Pseudomonas stutzeri WW88
-
-
-

Purification (Commentary)

Purification (Comment) Organism
on Ni2+ affinity resin Pseudomonas stutzeri

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
phosphite + H2O + NAD+ Asp79 is important in orienting Arg237 for proper interaction with phosphite Pseudomonas stutzeri phosphate + NADH + H+
-
?
phosphite + H2O + NAD+ Asp79 is important in orienting Arg237 for proper interaction with phosphite Pseudomonas stutzeri WW88 phosphate + NADH + H+
-
?

Synonyms

Synonyms Comment Organism
phosphite dehydrogenase
-
Pseudomonas stutzeri
PTDH
-
Pseudomonas stutzeri

Turnover Number [1/s]

Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
0.0126
-
phosphite mutant R237K Pseudomonas stutzeri
0.0305
-
phosphite mutant D79A Pseudomonas stutzeri
0.23
-
phosphite mutant D79N Pseudomonas stutzeri
0.58
-
phosphite mutant NADP-12X NAD Pseudomonas stutzeri
0.621
-
phosphite mutant K76A Pseudomonas stutzeri
2.04
-
phosphite mutant NADP-12X NAD Pseudomonas stutzeri
2.42
-
phosphite wild-type Pseudomonas stutzeri
3.26
-
phosphite thermostable mutant 12X-PTDH Pseudomonas stutzeri
7.5
-
phosphite mutant E266Q Pseudomonas stutzeri

Cofactor

Cofactor Comment Organism Structure
NAD(P)+
-
Pseudomonas stutzeri