High-performance liquid chromatography-electrospray ionization-mass spectrometry ligand fishing assay: a method for screening triplex DNA binders from natural plant extracts.

A novel ligand fishing assay was established to screen triplex DNA binders from complicated samples by a combination of immobilization of triplex DNA on agarose beads and high-performance liquid chromatography-electrospray ionization-mass spectrometry (HPLC-ESI-MS). The biotinylated oligodeoxynucleotides were first bound to the streptavidin agarose beads and then incubated with the duplex DNA as the baits for ligand fishing. This assay was validated by the testing ligand library consisting of coralyne, ethidium bromide, vitexin, and formononetin. The binding affinities of ligands to target DNA were also obtained based on the calibration curves of ligands. Two components (berberine and palmatine) in the extract of Phellodendron chinense Schneid cortexes were fished out as triplex DNA binders by this assay, which indicated its feasibility for screening triplex DNA binders from complicated samples. This preliminary assay can be used for not only screening binders of triplex DNA from natural products extracts but also can obtain their binding affinity information.