5'-Atp → 2-(6-Hydroxy-3-oxo-3H-xanthen-9-yl)benzoic acid

HRID 969254

Reaction equation

HRID 969254 reaction equation
C1N=C(N)C2N=CN([C@@H]3O[C@@H]4COP(O)(O[C@H]4[C@H]3O)=O)C=2N=1.P(OC[C@H]1O[C@@H](N2C3N=CN=C(N)C=3N=C2)[C@H](O)[C@@H]1O)(OP(OP(O)(O)=O)(O)=O)(=O)O.[Mg+2].[Cl-].[Cl-].C(S)[C@@H](O)[C@H](O)CS.[CH:65]1[CH:66]=[CH:67][C:68]([C:87]([OH:89])=[O:88])=[C:69]([C:71]2[C:81]3[CH:82]=[CH:83][C:84]([OH:86])=[CH:85][C:80]=3[O:79][C:78]3[C:72]=2[CH:73]=[CH:74][C:75]([CH:77]=3)=[O:76])[CH:70]=1.C[C@H](NC([C@@H](NC([C@@H](NC([C@@H](N)CC(C)C)=O)CCCNC(N)=N)=O)CCCNC(N)=N)=O)C(N[C@H](C(N[C@H](C(NCC(O)=O)=O)CC(C)C)=O)CO)=O.C[C@H](NC([C@@H](NC([C@@H](NC([C@@H](N)CC(C)C)=O)CCCNC(N)=N)=O)CCCNC(N)=N)=O)C(N[C@H](C(N[C@H](C(NCC(O)=O)=O)CC(C)C)=O)CO)=O>C1N(CCO)CCN(CCS(O)(=O)=O)C1>[CH:65]1[CH:66]=[CH:67][C:68]([C:87]([OH:89])=[O:88])=[C:69]([C:71]2[C:72]3[CH:73]=[CH:74][C:75]([OH:76])=[CH:77][C:78]=3[O:79][C:80]3[C:81]=2[CH:82]=[CH:83][C:84]([CH:85]=3)=[O:86])[CH:70]=1

Participants

8

Reactant 6

HCID 59575'-AtpHCID 6076CampHCID 446094D-1,4-DithiothreitolHCID 5360315Magnesium ChlorideHCID 178446007HCID 178446007, compoundHCID 178446008HCID 178446008, compound

Product 1

HCID 33832-(6-Hydroxy-3-oxo-3H-xanthen-9-yl)benzoic acid

Reagents, catalysts & solvents 1

HCID 238314-(2-Hydroxyethyl)-1-piperazineethanesulfonic acidSolvent2×

Procedure

PKA was diluted in 20 mM HEPES, pH 7.4, w/0.05% TX-100 and incubated in a final reaction buffer consisting of 20 mM HEPES, pH 7.4, 0.1 mM cAMP, 1 mM ATP, 5 mM MgCl2, 1 mM DTT with either fluorescein Kemptide (71.25 uM) or Oregon Green Kemptide as the enzymatic substrate at a final concentration of 71.25 or 114.25 μM, respectively, with a final reaction volume of 30 μl, for a reaction time of 1 hour in a 96 well plate format using an opaque white plate. Subsequently, 120 μl of 0.25×Fe(III) working reagent prepared according to Example I was added to the enzymatic reaction mixture and the fluorescence was measured using a 485/538 ex/em filter set. The results are shown in FIGS. 14 and 15.

Workup

  1. customreaction
  2. customrespectively, with a final reaction volume of 30 μl
  3. customfor a reaction time of 1 hour
  4. additionwas added to the enzymatic reaction mixture
  5. filtrationfilter

Sources & evidence

ORD record
ord-e3e3a1d134e94e878c394c49a6c30e47
Source dataset
uspto-grants-2007_12
Patent number
US07306928B2

Linked notes

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