Glucose oxidase as a model enzyme for antidiabetic activity evaluation of medicinal plants: In vitro and in silico evidences

Abstract

Background and objectives: Diabetes mellitus is a major public health problem in the world. In Africa, more than 80% of patients use plants for their treatment. However, the methods of validation of endogenous knowledge usually used are costly. The alternative method we propose aims at creating hyperglycemia in vitro and exploiting the metabolic pathway involving glucose oxidase for UV-visible spectrophotometric screening of the antidiabetic activity of medicinal plants.Methodology: The evolution of glucose oxidation as a function of drug concentration is followed by UV-visible spectrophotometry. The formation of the stable complex between the enzyme and the inhibitor is studied by molecular docking.Results: Drugs used (Gliben and plant extracts) displayed an in vitro hypoglycemic effect by exponentially reducing the level of free glucose in vitro. L. multiflora is more active than V. amygdalina (IC50: 1.36 ± 0.09 mg/mL Vs IC50: 3.00 ± 0.54 mg/mL). Gliben (0.5 mg/mL) and L. multiflora (2 mg/mL) reduced both the rate of oxidation of glucose by glucose oxidase (catalytic power Vmax: 0.84 ± 0.11 mg*mL-1*min-1 for Gliben and 1.72 ± 0.13 mg*mL-1*min-1 for L. multiflora) and the affinity of this enzyme for its substrate-glucose (KM: 15.11 ± 2.72 mg*mL-1 for Gliben and 9.17 ± 1.56 mg*mL-1 for L. multiflora) when compared to enzyme catalysis in the  absence of inhibitor (Vmax: 2.86 ± 0.44 mg*mL-1*min-1; KM: 8.07 ± 1.96 mg*mL-1). The binding of GOX (1GAL) to selected phyto-compounds derived from L. multiflora was confirmed by molecular docking. The best stability complexes were obtained for four compounds 8 (-10.3 Kcal/mol), 9 (-9.7 Kcal/mol), 6 (-9.7 Kcal/mol) and 3 (-9.5 Kcal/mol). Among these compounds, compounds 8 et 6 formed their complexes through  hydrogen bonds: the compound 8 form 04 hydrogen bond (ALA 292, GLY 290, SER 291, TYR 80) while the compound 6 form 03 hydrogen bonds (SER 103, THR 110, TYR 515). However, none H-bonding interaction occurs in the complex that involves ligands 9 and 3 despite their high binding energy (-9.7 Kcal/mol and -9.5 Kcal/mol respectively).Conclusion: Glucose oxidase can serve as a marker enzyme for in vitro antidiabetic activity evaluation of medicinal plants