Identification and characterization of dihydropyrimidinase inhibited by plumbagin isolated from Nepenthes miranda extract.
Dihydropyrimidinase is a member of the cyclic amidohydrolase family, which also includes allantoinase, dihydroorotase, hydantoinase, and imidase. This enzyme is important in pyrimidine metabolism, and blocking its activity would be detrimental to cell survival. This study investigated the dihydropyrimidinase inhibition by plumbagin isolated from the extract of carnivorous plant Nepenthes miranda (Nm). Plumbagin inhibited dihydropyrimidinase with IC50 value of 58 ± 3 μM. Double reciprocal results of Lineweaver-Burk plot indicated that this compound is a competitive inhibitor of dihydropyrimidinase. Fluorescence quenching analysis revealed that plumbagin could form a stable complex with dihydropyrimidinase with the Kd value of 37.7 ± 1.4 μM. Docking experiments revealed that the dynamic loop crucial for stabilization of the intermediate state in dihydropyrimidinase might be involved in the inhibition effect of plumbagin. Mutation at either Y155 or K156 within the dynamic loop of dihydropyrimidinase caused low plumbagin binding affinity. In addition to their dihydropyrimidinase inhibition, plumbagin and Nm extracts also exhibited cytotoxicity on melanoma cell survival, migration, and proliferation. Further research can directly focus on designing compounds that target the dynamic loop in dihydropyrimidinase during catalysis.
['Huang|Yen-Hua|YH|', 'Lien|Yi|Y|', 'Chen|Jung-Hung|JH|', 'Lin|En-Shyh|ES|', 'Huang|Cheng-Yang|CY|']
D000581:Amidohydrolases D000818:Animals D000900:Anti-Bacterial Agents D000970:Antineoplastic Agents D000975:Antioxidants D001419:Bacteria D002460:Cell Line D045744:Cell Line, Tumor D019684:Magnoliopsida D051379:Mice D009285:Naphthoquinones D010936:Plant Extracts
Dihydropyrimidinase Plumbagin Nepenthes miranda Allantoinase Dihydroorotase Imidase