title: Development of receptor-based inhibitory RNA aptamers for anthrax toxin neutralization.
abstract: Anthrax toxin excreted by Bacillus anthracis is the key causative agent of infectious anthrax disease. In the present study, we targeted the binding of PA to the ATR/TEM8 Von Willebrand factor type A (VWA) domain, which we cloned into Escherichia coli and purified to homogeneity under denaturing conditions. To develop an anthrax toxin inhibitor, we selected and identified short single strand RNA aptamers (approximately 30mer) consisting of different sequences of nucleic acids with a high binding affinity in the 100 nanomolar range against the recombinant ATR/TEM8 VWA domain using systematic evolution of ligands by exponential enrichment (SELEX). Five candidate aptamers were further characterized by several techniques including secondary structural analysis. The inhibitor efficiency (IC50) of one of the aptamers toward anthrax toxin was approximately 5μM in macrophage RAW 264.7 cells, as determined from cytotoxicity analysis by MTT assay. We believe that the candidate aptamers should be useful for blocking the binding of PA to its receptor in order to neutralize anthrax toxin.
authors.affiliation: Department of Chemistry and Research Institute of Natural Sciences, Hanyang University, Seoul 133-791, Republic of Korea.
authors.forename: Sang-Choon
authors.initials: SC
authors.lastname: Lee
authors.affiliation: Department of Chemistry and Research Institute of Natural Sciences, Hanyang University, Seoul 133-791, Republic of Korea.
authors.forename: Vinayakumar
authors.initials: V
authors.lastname: Gedi
authors.affiliation: Department of Chemistry and Research Institute of Natural Sciences, Hanyang University, Seoul 133-791, Republic of Korea.
authors.forename: Na-Reum
authors.initials: NR
authors.lastname: Ha
authors.affiliation: Department of Chemistry and Research Institute of Natural Sciences, Hanyang University, Seoul 133-791, Republic of Korea.
authors.forename: Jun-Haeng
authors.initials: JH
authors.lastname: Cho
authors.affiliation: Veterinary Drugs & Biologics Division, Animal and Plant Quarantine Agency (QIA), Anyang 430-757, Republic of Korea.
authors.forename: Hae-Chul
authors.initials: HC
authors.lastname: Park
authors.affiliation: Department of Chemistry and Research Institute of Natural Sciences, Hanyang University, Seoul 133-791, Republic of Korea. Electronic address: myyoon@hanyang.ac.kr.
authors.forename: Moon-Young
authors.initials: MY
authors.lastname: Yoon
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D000942:Antigens, Bacterial / Q000378:metabolism*; D052157:Aptamers, Nucleotide / Q000378:metabolism* / Q000633:toxicity; D001427:Bacterial Toxins / Q000378:metabolism*; D006801:Humans; D051379:Mice; D008840:Microfilament Proteins; D008969:Molecular Sequence Data; D009363:Neoplasm Proteins / Q000737:chemistry / Q000378:metabolism*; D017434:Protein Structure, Tertiary; D000067996:RAW 264.7 Cells; D011956:Receptors, Cell Surface / Q000737:chemistry / Q000378:metabolism*; D052156:SELEX Aptamer Technique
chemical_list: C473287:ANTXR1 protein, human; D000942:Antigens, Bacterial; D052157:Aptamers, Nucleotide; D001427:Bacterial Toxins; D008840:Microfilament Proteins; D009363:Neoplasm Proteins; D011956:Receptors, Cell Surface; C030325:anthrax toxin
keywords: ATR; Anthrax toxin; RNA aptamer