Targeting of Single‐Stranded Oligonucleotides through Metal‐Induced Cyclization of Short Complementary Strands

Abstract

A new strategy to cyclize short synthetic oligonucleotides on DNA or RNA target strands is described. The approach is based on metal‐templated cyclization of short synthetic oligonucleotides conjugated with two chelating 2,2′ : 6′,2′′‐terpyridine (Tpy) moieties at their 3′‐ and 5′‐ends. Cyclization after metal addition (Zn2+, Fe2+) was demonstrated by means of thermal‐denaturation experiments, MALDI‐Q‐TOF‐MS, and gel electrophoresis (PAGE). 1D‐ and 2D‐NMR Experiments were performed to analyze the association of complementary strands after metal‐mediated cyclization. Our protocol allows the efficient circularization of synthetic oligonucleotides. Thereby, the hybridization on a complementary strand was more efficient with an RNA target strand and a 2′‐O‐methylated circularized oligomer.

Reversible Metal-Dependent Destabilization and Stabilization of a Stem-Chelate-Loop Probe Binding to an Unmodified DNA Target

“…We previously reported the development of PNA-based probes that exhibited enhanced binding specificity. 23 In an analogous arrangement, Krämer 24 and Moreau 21 introduced oligonucleotides with antipodal TPY groups that underwent macrocyclization in the presence of a coordinating transition metal. Chelate-modified oligonucleotides consisting of unnatural metallo-base pairs ( e.g.…”

“…In control transitions absent free metal ions (i.e., excess EDTA), TPY-containing probe–target duplexes 4:10 and 6:10 were both stabilized by ∼2 °C relative to their TPY-free counterparts (Figure , gray segment). TPY-base-pair stacking interactions provide a duplex stabilizing force, particularly with 3′ adjacent purine nucleobases …”

“…We previously reported the development of PNA-based probes that exhibited enhanced binding specificity . In an analogous arrangement, Krämer and Moreau introduced oligonucleotides with antipodal TPY groups that underwent macrocyclization in the presence of a coordinating transition metal. Chelate-modified oligonucleotides consisting of unnatural metallo-base pairs (e.g., T-Hg + -T) are known that exhibit exceptional duplex stability. − However, to our knowledge this is the first report of a novel stem-chelate-loop nucleobase polymer assembly that permits metal-dependent stabilization and destabilization of probe-target binding to unmodified DNA.…”