title: Dimerization and DNA recognition rules of mithramycin and its analogues.
abstract: The antineoplastic and antibiotic natural product mithramycin (MTM) is used against cancer-related hypercalcemia and, experimentally, against Ewing sarcoma and lung cancers. MTM exerts its cytotoxic effect by binding DNA as a divalent metal ion (Me(2+))-coordinated dimer and disrupting the function of transcription factors. A precise molecular mechanism of action of MTM, needed to develop MTM analogues selective against desired transcription factors, is lacking. Although it is known that MTM binds G/C-rich DNA, the exact DNA recognition rules that would allow one to map MTM binding sites remain incompletely understood. Towards this goal, we quantitatively investigated dimerization of MTM and several of its analogues, MTM SDK (for Short side chain, DiKeto), MTM SA-Trp (for Short side chain and Acid), MTM SA-Ala, and a biosynthetic precursor premithramycin B (PreMTM B), and measured the binding affinities of these molecules to DNA oligomers of different sequences and structural forms at physiological salt concentrations. We show that MTM and its analogues form stable dimers even in the absence of DNA. All molecules, except for PreMTM B, can bind DNA with the following rank order of affinities (strong to weak): MTM=MTM SDK>MTM SA-Trp>MTM SA-Ala. An X(G/C)(G/C)X motif, where X is any base, is necessary and sufficient for MTM binding to DNA, without a strong dependence on DNA conformation. These recognition rules will aid in mapping MTM sites across different promoters towards development of MTM analogues as useful anticancer agents.
authors.affiliation: Department of Pharmaceutical Sciences, College of Pharmacy, University of Kentucky, Lexington, KY 40536, USA.
authors.forename: Stevi
authors.initials: S
authors.lastname: Weidenbach
authors.affiliation: Department of Pharmaceutical Sciences, College of Pharmacy, University of Kentucky, Lexington, KY 40536, USA.
authors.forename: Caixia
authors.initials: C
authors.lastname: Hou
authors.affiliation: Department of Pharmaceutical Sciences, College of Pharmacy, University of Kentucky, Lexington, KY 40536, USA.
authors.forename: Jhong-Min
authors.initials: JM
authors.lastname: Chen
authors.affiliation: Department of Pharmaceutical Sciences, College of Pharmacy, University of Kentucky, Lexington, KY 40536, USA. Electronic address: oleg.tsodikov@uky.edu.
authors.forename: Oleg V
authors.initials: OV
authors.lastname: Tsodikov
authors.affiliation: Department of Pharmaceutical Sciences, College of Pharmacy, University of Kentucky, Lexington, KY 40536, USA. Electronic address: jrohr2@uky.edu.
authors.forename: Jürgen
authors.initials: J
authors.lastname: Rohr
mesh_terms: D000903:Antibiotics, Antineoplastic / Q000737:chemistry*; D004247:DNA / Q000737:chemistry*; D019281:Dimerization; D008926:Plicamycin / Q000737:chemistry*
chemical_list: D000903:Antibiotics, Antineoplastic; D004247:DNA; D008926:Plicamycin
keywords: Anticancer agent; DNA binding; Metal ion coordination; Minor groove; Natural product