Reaction #1755459

ord-01cba8bec4c749768425947ff25e45f9

Reaction equation

Cl.NC(CO)(CO)CO
Tris HCl
NC(=O)c1ccc[n+]([C@@H]2O[C@H](COP(=O)(O)OP(=O)(O)OC[C@H]3O[C@@H](n4cnc5c(N)ncnc54)[C@H](OP(=O)(O)O)[C@@H]3O)[C@@H](O)[C@H]2O)c1
NADP
[Cl-].[Cl-].[Mg+2]
magnesium chloride
O=C(O)[C@H](O)[C@@H](O)[C@H](O)[C@H](O)COP(=O)(O)O
6-phosphogluconic acid
→
NC(=O)C1=CN([C@@H]2O[C@H](COP(=O)(O)OP(=O)(O)OC[C@H]3O[C@@H](n4cnc5c(N)ncnc54)[C@H](OP(=O)(O)O)[C@@H]3O)[C@@H](O)[C@H]2O)C=CC1
NADPH

Reactants

Cl.NC(CO)(CO)CO
Tris HCl
NC(=O)c1ccc[n+]([C@@H]2O[C@H](COP(=O)(O)OP(=O)(O)OC[C@H]3O[C@@H](n4cnc5c(N)ncnc54)[C@H](OP(=O)(O)O)[C@@H]3O)[C@@H](O)[C@H]2O)c1
NADP
[Cl-].[Cl-].[Mg+2]
magnesium chloride
O=C(O)[C@H](O)[C@@H](O)[C@H](O)[C@H](O)COP(=O)(O)O
6-phosphogluconic acid

Reagents

None

Conditions

Detailed conditions
See reaction.notes.procedure_details.

Workup

  1. 1
    OtherThe resulting mixture was subjected to reaction in a 1-ml cuvette at 30° C.
  2. 2
    Temperaturethe increase in the absorbance

Procedure

The crude enzyme solution was added to a reaction mixture comprising 50 mmol/l Tris/HCl (pH 7.5), 0.5 mmol/l NADP, 10 mmol/l magnesium chloride and 2 mmol/l 6-phosphogluconic acid to make up to 1.5 ml. The resulting mixture was subjected to reaction in a 1-ml cuvette at 30° C. and the increase in the absorbance was measured at 340 nm. On the basis of the molar extinction coefficient of NADPH (6220), the amount of NADPH formed per minute was calculated. GND specific activity was expressed in terms of the amount of NADPH formed per mg of protein per minute.

Source

DOI: 10.6084/m9.figshare.5104873.v1Patent: US07989191B2uspto-grants-2011_08