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In this study, we characterized the enzymology of cytosolic catechol-O-methyltransferase (COMT)-catalyzed …


Biology Articles » Biochemistry » Enzymology » Enzymology of Methylation of Tea Catechins and Inhibition of Catechol-O-methyltransferase by ()-Epigallocatechin Gallate » Tables

Tables
- Enzymology of Methylation of Tea Catechins and Inhibition of Catechol-O-methyltransferase by ()-Epigallocatechin Gallate

TABLE 1
Kinetic parameters of EGCG and EGC methylation by liver cytosolic COMT

Kinetic studies were conducted for 2 min (EGCG) or 5 min (EGC) using 1 mg/ml of liver cytosolic proteins in the presence of 50 µM SAM. The kinetic parameters (Km and Vmax) were calculated from duplicated determinations with GraphPad Prism. Values represent best-fit values ± standard error.

EGCG EGC
Vmax Km Vmax/Km Vmax Km Vmax/Km
Human liver 161  ± 4 0.16  ± 0.02 1000 1284  ± 86 4.0  ± 1.4 321
Mouse liver 145  ± 7 0.17  ± 0.05 853 757  ± 21 8.1  ± 0.9 93
Rat liver 413  ± 21 0.48  ± 0.10 860 2130  ± 57 11.7  ± 1.0 182
Vmax, picomoles per milligram per minute; Km, micromolar; Vmax/Km, microliter per milligram per minute.

TABLE 2
Median inhibition concentration (IC50) of the inhibition of methylation of EGC and L-DOPA by catechins and their derivatives

Studies on the inhibition of EGC methylation were conducted for 10 min using 1 mg/ml of liver cytosolic proteins in the presence of 10 µM EGC, 50 µM SAM and different concentrations of inhibitors. Studies on the inhibition of L-DOPA methylation were conducted for 30 min using 25 units of porcine liver COMT in the presence of 100 µM L-DOPA, 60 µM SAM and different concentrations of inhibitors. Values of IC50 (µM) were calculated from duplicate determinations and represent the concentrations of inhibitor that inhibited the formation of 4'-MeEGC or 3-MeDOPA by 50%.

Inhibitor COMT Substrate Inhibitor COMT Substrate
EGC L-DOPA EGC L-DOPA
EGCG 0.15 0.20 ECG 0.25 0.90
4"-MeEGCG 0.16 0.10 EGC N/D 19
4',4"-DiMeEGCG 0.20 0.30 (-)-Epicatechin N/D 64
4',3",4"-TriMeEGCG 0.15 N/D 4'-MeEGC N/D >100
EGCG-7-Gluc 0.30 0.50 EGC-3'-Gluc N/D >100
EGCG-3'-Gluc 0.11 N/D EGC-7-Gluc N/D 16
EGCG-3"-Gluc 0.15 N/D 3,5-Dinitrocatechol 0.045 0.10
EGCG-4"-Gluc 1.5 2.5 Pyrogallol 9.0 N/D
N/D, not determined.


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