The present study aims to investigate the substrate specificity of peroxidase(POD),an enzyme held responsible for enzymatic browning,in the presence of caffeic acid,guaiacol,chlorogenic acid and catechol.The use of in...The present study aims to investigate the substrate specificity of peroxidase(POD),an enzyme held responsible for enzymatic browning,in the presence of caffeic acid,guaiacol,chlorogenic acid and catechol.The use of in silico studies provided additional support for the identification of optimal substrate candidates.Considering the Vmax/Km(EU/mL.min.mM)values obtained,caffeic acid was found to have the highest value(11.85)and to be the best substrate for POD.The inhibitory mechanism of ascorbic acid,citric acid and salicylic acid on the activity of POD was studied in the presence of caffeic acid.Competitive inhibition was observed in the kinetic studies involving ascorbic acid and salicylic acid,whereas non-competitive inhibition was observed in the case of citric acid.The lowest calculated dissociation constant(Ki)of the enzyme-inhibitor complex was that of salicylic acid(0.044 mM).Furthermore,the in vitro Ki results for the inhibitors used were consistent with the in silico studies.The mechanism of POD inhibition by salicylic acid in the presence of a caffeic acid substrate was explained by molecular docking studies.The POD inhibition has been attributed to a decrease in substrate oxidation as a result of ferric(H2O2)interactions with salicylic acid,in addition to a decrease in enzyme stabilization as a result of Pi-lone pair interactions.展开更多
文摘The present study aims to investigate the substrate specificity of peroxidase(POD),an enzyme held responsible for enzymatic browning,in the presence of caffeic acid,guaiacol,chlorogenic acid and catechol.The use of in silico studies provided additional support for the identification of optimal substrate candidates.Considering the Vmax/Km(EU/mL.min.mM)values obtained,caffeic acid was found to have the highest value(11.85)and to be the best substrate for POD.The inhibitory mechanism of ascorbic acid,citric acid and salicylic acid on the activity of POD was studied in the presence of caffeic acid.Competitive inhibition was observed in the kinetic studies involving ascorbic acid and salicylic acid,whereas non-competitive inhibition was observed in the case of citric acid.The lowest calculated dissociation constant(Ki)of the enzyme-inhibitor complex was that of salicylic acid(0.044 mM).Furthermore,the in vitro Ki results for the inhibitors used were consistent with the in silico studies.The mechanism of POD inhibition by salicylic acid in the presence of a caffeic acid substrate was explained by molecular docking studies.The POD inhibition has been attributed to a decrease in substrate oxidation as a result of ferric(H2O2)interactions with salicylic acid,in addition to a decrease in enzyme stabilization as a result of Pi-lone pair interactions.
