title: Competitive binding between 4,4'-diphenylmethane-bis(methyl) carbamate and RAGE ligand MG-H1 on human umbilical vein endothelial cell by cell membrane chromatography.
abstract: The compound 4,4'-diphenylmethane-bis(methyl) carbamate (CM1) has a protective activity on AGEs-induced endothelial dysfunction on human umbilical vein endothelial cell (HUVEC) in our previous study. It suggested that CM1 which may act as a competitive antagonist to the blockade of AGEs to receptor of AGEs (RAGE) and attenuate the HUVEC damage. In order to testify that hypothesis, the cell membrane chromatography (CMC) combined with high performance liquid chromatography (HPLC) was developed for analyzing the competitive binding properties on RAGE of HUVEC between CM1 and MG-H1, the agonist of RAGE. The results from saturation binding of CM1 and MG-H1 on cells demonstrated that dissociation equilibrium constants (K(d)) of CM1 and MG-H1 were 3.653 nM and 4.12 nM, respectively; while maximum binding capacity (B(max)) of CM1 and MG-H1 were 30.08 and 18.72 fmol/mg protein, respectively. In competition experiments, IC₅₀ of CM1 with pre-incubation 10⁻¹⁰ M and 10⁻⁹ M MG-H1 were 1.37 × 10⁻⁹ M and 4.56 × 10⁻⁸ M, respectively. The present findings indicated that CM1 conjugated competitively to cells with RAGE ligand MG-H1. The primary study illustrated that CMC combined with HPLC analysis method could be an alternative, rapid and efficient approach for the interaction of drug molecule and receptor, and that CM1 intervene the AGEs inducing HUVEC damage may via the competitively block the AGEs-RAGE path way.
authors.affiliation: State Key Laboratory of Quality Research in Chinese Medicine, Biotechnology Laboratory of Chinese Medicine, Macau University of Science and Technology, Taipa, Macau.
authors.forename: Liang
authors.initials: L
authors.lastname: Feng
authors.forename: You-Hua
authors.initials: YH
authors.lastname: Xu
authors.forename: Shan-Shan
authors.initials: SS
authors.lastname: Wang
authors.forename: Wai
authors.initials: W
authors.lastname: Au-Yeung
authors.forename: Zhao-Guang
authors.initials: ZG
authors.lastname: Zheng
authors.forename: Quan
authors.initials: Q
authors.lastname: Zhu
authors.forename: Ping
authors.initials: P
authors.lastname: Xiang
mesh_terms: D001667:Binding, Competitive; D002219:Carbamates / Q000737:chemistry / Q000378:metabolism*; D002462:Cell Membrane / Q000378:metabolism*; D002851:Chromatography, High Pressure Liquid; D002853:Chromatography, Liquid / Q000379:methods*; D017127:Glycation End Products, Advanced; D061307:Human Umbilical Vein Endothelial Cells / Q000378:metabolism*; D006801:Humans; D007700:Kinetics; D008024:Ligands; D009952:Ornithine / Q000031:analogs & derivatives* / Q000378:metabolism; D000067759:Receptor for Advanced Glycation End Products; D011971:Receptors, Immunologic / Q000819:agonists* / Q000378:metabolism
chemical_list: D002219:Carbamates; D017127:Glycation End Products, Advanced; D008024:Ligands; D000067759:Receptor for Advanced Glycation End Products; D011971:Receptors, Immunologic; D009952:Ornithine