title: High throughput screening of high-affinity ligands for proteins with anion-binding sites using desorption electrospray ionization (DESI) mass spectrometry.
abstract: A high throughput screening system involving a linear ion trap (LTQ) analyzer, a house-made platform and a desorption electrospray ionization (DESI) source was established to screen ligands with a high affinity for proteins with anion-binding sites. The complexes were analyzed after incubation, ultrafiltration, washing, and displacement. A new anionic region inhibited dissociation (ARID) mechanism that was suitable for a protein with anion-binding site was proposed. We utilized the differences in detectable dissociation of protein-ligand complexes, combined with displacement experiments, to distinguish free ligands displaced from anion-binding sites from liberated ligands dissociated from nonspecific interactions. The method was validated by α1-acid glycoprotein (AGP) and (R), (S)-amlodipine. Site-specific enantioselectivity shown in our experiments was consistent with earlier studies. Obtaining all of the qualitative information of 15*3 samples in 2.3 min indicates that the analysis process is no longer the time-limiting step in the initial stage of drug discovery. Quantitative information verified that our method was at least a semiquantitative method.
authors.affiliation: National Institutes for Food and Drug Control, Beijing, 100050, Peoples Republic of China.
authors.forename: Xin
authors.initials: X
authors.lastname: Lu
authors.forename: Baoming
authors.initials: B
authors.lastname: Ning
authors.forename: Dacheng
authors.initials: D
authors.lastname: He
authors.forename: Lingyun
authors.initials: L
authors.lastname: Huang
authors.forename: Xiangjun
authors.initials: X
authors.lastname: Yue
authors.forename: Qiming
authors.initials: Q
authors.lastname: Zhang
authors.forename: Haiwei
authors.initials: H
authors.lastname: Huang
authors.forename: Yang
authors.initials: Y
authors.lastname: Liu
authors.forename: Lan
authors.initials: L
authors.lastname: He
authors.forename: Jin
authors.initials: J
authors.lastname: Ouyang
mesh_terms: D000838:Anions / Q000032:analysis* / Q000737:chemistry / Q000378:metabolism*; D001665:Binding Sites; D057166:High-Throughput Screening Assays / Q000379:methods*; D008024:Ligands; D011485:Protein Binding; D011506:Proteins / Q000737:chemistry / Q000378:metabolism*; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods*
chemical_list: D000838:Anions; D008024:Ligands; D011506:Proteins