Bifacial Peptide Nucleic Acid as an Allosteric Switch for Aptamer and Ribozyme Function

Abstract

We demonstrate herein that bifacial peptide nucleic acid (bPNA) hybrid triplexes functionally substitute for duplex DNA or RNA. Structure-function loss in three non-coding nucleic acids was inflicted by replacement of a duplex stem with unstructured oligo-T/U strands, which are bPNA binding sites. Functional rescue was observed on refolding of the oligo-T/U strands into bPNA triplex hybrid stems. Bifacial PNA binding was thus used to allosterically switch-on protein and small-molecule binding in DNA and RNA aptamers, as well as catalytic bond scission in a ribozyme. Duplex stems that support the catalytic site of a minimal type I hammerhead ribozyme were replaced with oligo-U loops, severely crippling or ablating the native RNA splicing function. Refolding of the U-loops into bPNA triplex stems completely restored splicing function in the hybrid system. These studies indicate that bPNA may have general utility as an allosteric trigger for a wide range of functions in non-coding nucleic acids.

Was a Pyrimidine‐Pyrimidine Base Pair the Ancestor of Watson‐Crick Base Pairs? Insights from a Systematic Approach to the Origin of RNA

“…Eschenmoser and coworkers have shown that peptide nucleic acid backbones tagged with complementary triazines can, albeit weakly, form homo‐duplexes and cross‐pair to form more thermally stable hetero‐duplexes with both DNA and RNA 66. The Bong laboratory has also shown that peptide nucleic acids tagged with melamine (the triazine analog of TAP) are able to form triplex assemblies with either pol‐U or poly‐dT,67 demonstrating melamine’s compatibility with the extant nucleobases.…”

A modular clamp-like mechanism to regulate the activity of nucleic-acid target-responsive nanoswitches with external activators

“…Pioneer works from the groups of Breaker and Famulok have recreated in-vitro allosteric regulation to control ribozymes 52 and nucleic acid target-responsive elements. 53 Other applications of allosteric regulation include sensing [53][54][55][56][57] and those in the emerging field of synthetic biology. [58][59] Here we have expanded on this theme and have rationally designed novel clamp-like target-responsive DNA-based nanoswitches that, by using a strategy similar to that of allosterically controlled intrinsically disordered proteins, can be activated by a partial folding induced by the binding of an external activator.…”

“…Compared to other methods where modulation of the activity of DNA-based receptors [54][55][56][57] has been achieved with allosteric approaches our strategy appears particularly advantageous for several reasons. Because it takes advantage of a triplex-forming nucleic acid recognition module that can be triggered in a highly specific and sensitive way by natural DNA strands, 44 our approach appears far more suitable for sensing and synthetic biology applications than those based on triplex-forming portions triggered by non-natural targets (i.e.…”

Molecular Glue for RNA: Regulating RNA Structure and Function through Synthetic RNA Binding Molecules

“…[17] These findings indicate that the activity of HHR could be modulated by am olecular glue that brings the two loops into contact to form the higher-order structureo fH HR. [15,22] To construct a( Z)-NCTS-inducible HHR, we modifiedl oops I and II by replacing fiveb ases, therebyd isrupting the looploop interaction ( Figure 6). [15] An HHR lacking this tertiaryc ontact loses the ability to cleave RNA ( Figure 6B).…”