title: Aptamer selection based on G4-forming promoter region.
abstract: We developed a method for aptamer identification without in vitro selection. We have previously obtained several aptamers, which may fold into the G-quadruplex (G4) structure, against target proteins; therefore, we hypothesized that the G4 structure would be an excellent scaffold for aptamers to recognize the target protein. Moreover, the G4-forming sequence contained in the promoter region of insulin can reportedly bind to insulin. We thus expected that G4 DNAs, which are contained in promoter regions, could act as DNA aptamers against their gene products. We designated this aptamer identification method as "G4 promoter-derived aptamer selection (G4PAS)." Using G4PAS, we identified vascular endothelial growth factor (VEGF)165, platelet-derived growth factor-AA (PDGF)-AA, and RB1 DNA aptamers. Surface plasmon resonance (SPR) analysis revealed that the dissociation constant (K d) values of VEGF165, PDGF-AA, and RB1 DNA aptamers were 1.7 × 10(-7) M, 6.3 × 10(-9) M, and 4.4 × 10(-7) M, respectively. G4PAS is a simple and rapid method of aptamer identification because it involves only binding analysis of G4 DNAs to the target protein. In the human genome, over 40% of promoters contain one or more potential G4 DNAs. G4PAS could therefore be applied to identify aptamers against target proteins that contain G4 DNAs on their promoters.
authors.affiliation: Department of Biotechnology and Life Science, Tokyo University of Agriculture & Technology, Koganei, Tokyo, Japan.
authors.forename: Wataru
authors.initials: W
authors.lastname: Yoshida
authors.forename: Taiki
authors.initials: T
authors.lastname: Saito
authors.forename: Tomomi
authors.initials: T
authors.lastname: Yokoyama
authors.forename: Stefano
authors.initials: S
authors.lastname: Ferri
authors.forename: Kazunori
authors.initials: K
authors.lastname: Ikebukuro
mesh_terms: D052157:Aptamers, Nucleotide / Q000378:metabolism*; D004247:DNA / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D054856:G-Quadruplexes; D006801:Humans; D007700:Kinetics; D010982:Platelet-Derived Growth Factor / Q000235:genetics / Q000378:metabolism; D011401:Promoter Regions, Genetic; D019009:Proto-Oncogene Proteins c-kit / Q000235:genetics / Q000378:metabolism; D016160:Retinoblastoma Protein / Q000235:genetics / Q000378:metabolism; D052156:SELEX Aptamer Technique / Q000379:methods*; D013379:Substrate Specificity; D042461:Vascular Endothelial Growth Factor A / Q000235:genetics / Q000378:metabolism
chemical_list: D052157:Aptamers, Nucleotide; D010982:Platelet-Derived Growth Factor; D016160:Retinoblastoma Protein; C467484:VEGFA protein, human; D042461:Vascular Endothelial Growth Factor A; C075278:platelet-derived growth factor A; D004247:DNA; D019009:Proto-Oncogene Proteins c-kit