title: Reversible metal-dependent destabilization and stabilization of a stem-chelate-loop probe binding to an unmodified DNA target.
abstract: Herein, we report the discovery of a novel DNA probe with a stem-chelate-loop structure, wherein the stability of the probe-target duplex can be modulated lower or higher using a narrow concentration range of dilute transition metal ions (0.1-10 μM). Oligonucleotide probes containing two terpyridine (TPY) ligands separated by 15 bases of single-stranded DNA, with or without a flanking 5 base self-complementary DNA stem, were tested in thermal transition studies with linear target DNA and varying amounts of ZnCl(2). Without the stem, addition of Zn(2+) resulted only in reversible destabilization of the probe-target duplex, consistent with assembly (up to 1 equiv Zn(2+)) and disassembly (excess Zn(2+)) of the intramolecular Zn(2+)-(TPY)(2) chelate. Surprisingly, probes including both the intramolecular chelate and the stem gave a probe-target duplex that was reversibly destabilized and stabilized upon addition of Zn(2+) by ±5-7 °C, a phenomenon consistent with assembly and then disassembly of the entire stem-Zn(2+)-(TPY)(2) motif, including the DNA stem. Stem-chelate-loop probes containing dipicolylamine (DPA) ligands exhibited no metal-dependent stabilization or destabilization. The stem-Zn(2+)-(TPY)(2) motif is readily introduced with automated synthesis, and may have broad utility in applications where it is desirable to have both upward and downward, reversible metal-dependent control over probe-target stability involving an unmodified DNA target.
authors.affiliation: Syntrix Biosystems, Inc., 215 Clay Street NW, Suite B-5, Auburn, WA 98001, USA. jmorgan@syntrixbio.com
authors.forename: Joel R
authors.initials: JR
authors.lastname: Morgan
authors.forename: David V X
authors.initials: DV
authors.lastname: Nguyen
authors.forename: Angela R
authors.initials: AR
authors.lastname: Frohman
authors.forename: Sara R
authors.initials: SR
authors.lastname: Rybka
authors.forename: John A
authors.initials: JA
authors.lastname: Zebala
mesh_terms: D001483:Base Sequence; D002614:Chelating Agents / Q000737:chemistry*; D004247:DNA / Q000737:chemistry* / Q000235:genetics; D015342:DNA Probes / Q000737:chemistry* / Q000235:genetics; D055029:Inverted Repeat Sequences / Q000235:genetics; D008958:Models, Molecular; D009690:Nucleic Acid Conformation; D009693:Nucleic Acid Hybridization; D015032:Zinc / Q000737:chemistry*
chemical_list: D002614:Chelating Agents; D015342:DNA Probes; D004247:DNA; D015032:Zinc