title: Electrophoretic behavior of DNA-methyl-CpG-binding domain protein complexes revealed by capillary electrophoreses laser-induced fluorescence.
abstract: The free solution electrophoretic behavior of DNA-protein complexes depends on their charge and mass in a certain experimental condition, which are two fundamental properties of DNA-protein complexes in free solution. Here, we used CE LIF to study the free solution behavior of DNA-methyl-CpG-binding domain protein (MBD2b) complexes through exploring the relationship between the mobilities, charge, and mass of DNA-protein complexes. This method is based on the effective separation of free DNA and DNA-protein complexes because of their different electrophoretic mobility in a certain electric field. In order to avoid protein adsorption, a polyacrylamide-coated capillary was used. Based on the evaluation of the electrophoretic behavior of formed DNA-MBD2b complexes, we found that the values of (μ0 /μ)-1 were directly proportional to the charge-to-mass ratios of formed complexes, where the μ0 and μ are the mobility of free DNA probe and DNA-protein complex, respectively. The models were further validated by the complex mobilities of protein with various lengths of DNA probes. The deviation of experimental and calculated charge-to-mass ratios of formed complexes from the theoretical data was less than 10%, suggesting that our models are useful to analyze the DNA-binding properties of the purified MBD2b protein and help to analyze other DNA-protein complexes. Additionally, this study enhances the understanding of the influence of the charge-to-mass ratios of formed DNA-protein complexes on their separation and electrophoretic behaviors.
authors.affiliation: College of Chemistry and Chemical Engineering, University of Chinese Academy of Sciences, Beijing, P. R. China.
authors.forename: Shangwei
authors.initials: S
authors.lastname: Zhong
authors.affiliation: State Key Laboratory of Environmental Chemistry and Ecotoxicology, Research Center for Eco-Environmental Sciences, Chinese Academy of Sciences, Beijing, P. R. China.
authors.forename: Dandan
authors.initials: D
authors.lastname: Zou
authors.affiliation: State Key Laboratory of Environmental Chemistry and Ecotoxicology, Research Center for Eco-Environmental Sciences, Chinese Academy of Sciences, Beijing, P. R. China.
authors.forename: Bailin
authors.initials: B
authors.lastname: Zhao
authors.affiliation: State Key Laboratory of Environmental Chemistry and Ecotoxicology, Research Center for Eco-Environmental Sciences, Chinese Academy of Sciences, Beijing, P. R. China.
authors.forename: Dapeng
authors.initials: D
authors.lastname: Zhang
authors.affiliation: College of Chemistry and Chemical Engineering, University of Chinese Academy of Sciences, Beijing, P. R. China.
authors.forename: Xiangjun
authors.initials: X
authors.lastname: Li
authors.affiliation: State Key Laboratory of Environmental Chemistry and Ecotoxicology, Research Center for Eco-Environmental Sciences, Chinese Academy of Sciences, Beijing, P. R. China.
authors.forename: Hailin
authors.initials: H
authors.lastname: Wang
mesh_terms: D004247:DNA / Q000032:analysis / Q000737:chemistry*; D004268:DNA-Binding Proteins / Q000032:analysis / Q000737:chemistry*; D019075:Electrophoresis, Capillary / Q000379:methods*; D005453:Fluorescence; D006801:Humans; D008970:Molecular Weight; D046912:Multiprotein Complexes / Q000032:analysis / Q000737:chemistry*; D011485:Protein Binding; D015203:Reproducibility of Results
chemical_list: D004268:DNA-Binding Proteins; C115368:MBD2 protein; D046912:Multiprotein Complexes; D004247:DNA
keywords: CE LIF; Charge-to-mass ratios; DNA-methyl-CpG binding domain protein; DNA-protein complexes; Electrophoretic behavior