title: Specificity and inhibitory mechanism of andrographolide and its analogues as antiasthma agents on NF-κB p50.
abstract: Andrographolide (1) is a diterpenoid lactone with an α,β-unsaturated lactone group that inhibits NF-κB DNA binding. Andrographolide reacts with the nucleophilic Cys62 of NF-κB p50 through a Michael addition at the Δ(12(13)) exocylic double bond to form a covalent adduct. Using computer docking, site-directed mutagenesis, and mass spectrometry, the noncovalent interactions between andrographolide and additional binding site residues other than Cys62 were found to be essential for the covalent incorporation of andrographolide. Furthermore, the addition reaction of andrographolide on Cys62 was highly dependent on the redox conditions and on the vicinity of nearby, positively charged Arg residues in the conserved RxxRxR motif. The reaction mechanisms of several of the analogues were determined, showing that 14-deoxy-11,12-didehydroandrographolide (8) reacts with NF-κB p50 via a novel mechanism distinct from andrographolide. The noncovalent interaction and redox environment of the binding site should be considered, in addition to the electrophilicity, when designing a covalent drug. Analogues similar in structure appear to use distinct reaction mechanisms and may have very different cytotoxicities, e.g., compound 6.
authors.affiliation: Department of Biological Sciences, National University of Singapore , 14 Science Drive 4, Singapore 117543.
authors.forename: Van Sang
authors.initials: VS
authors.lastname: Nguyen
authors.forename: Xin Yi
authors.initials: XY
authors.lastname: Loh
authors.forename: Hadhi
authors.initials: H
authors.lastname: Wijaya
authors.forename: Jigang
authors.initials: J
authors.lastname: Wang
authors.forename: Qingsong
authors.initials: Q
authors.lastname: Lin
authors.forename: Yulin
authors.initials: Y
authors.lastname: Lam
authors.forename: Wai-Shiu Fred
authors.initials: WS
authors.lastname: Wong
authors.forename: Yu Keung
authors.initials: YK
authors.lastname: Mok
mesh_terms: D035685:Andrographis / Q000737:chemistry*; D018927:Anti-Asthmatic Agents / Q000737:chemistry / Q000494:pharmacology*; D003545:Cysteine / Q000737:chemistry; D004224:Diterpenes / Q000737:chemistry / Q000494:pharmacology*; D015394:Molecular Structure; D016328:NF-kappa B / Q000037:antagonists & inhibitors*; D010084:Oxidation-Reduction
chemical_list: C495626:14-deoxy-11,12-didehydroandrographolide; D018927:Anti-Asthmatic Agents; D004224:Diterpenes; D016328:NF-kappa B; C030419:andrographolide; D003545:Cysteine