Divalent Nucleobases for RNA Mismatch Targeting
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Solution Overview
Problem
Current methods lack effective tools to specifically target and manipulate RNA structures, particularly mismatched sequences, which are crucial for understanding and modulating RNA functions and treating genetic and infectious diseases.
Innovation Solution
Development of divalent nucleobases that can form directional hydrogen bonding interactions with two strands of DNA or RNA, regardless of mismatches, allowing for specific binding and modulation of RNA interactions.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Reliability
If standard Watson-Crick base-pairing rules are used for targeting nucleic acids, then binding specificity to canonical sequences is achieved, but inability to effectively target mismatched sequences occurs
Solution Approach 1:
The invention divides the base-pairing interaction into two independent strands, each recognizing one side of the mismatch. The divalent nucleobase consists of two monovalent nucleobases linked by a connector, allowing each nucleobase to independently form Watson-Crick or wobble base pairs with complementary bases on opposite strands, thereby enabling specific recognition of mismatched sequences while maintaining binding reliability
Solution Approach 2:
The divalent nucleobase is constructed as a composite structure combining two monovalent nucleobases (adenine, guanine, cytosine, or uracil) with a connector molecule. This composite design allows the molecule to simultaneously engage both strands of the nucleic acid duplex at the mismatch site, providing both the stability of canonical base-pairing and the versatility to target non-canonical sequences
2Reliability
If proteins are used to target genetic information, then high binding affinity and specificity can be achieved, but complexity of the molecular tool increases
Solution Approach 1:
The invention changes the fundamental parameter of base-pairing geometry by introducing a divalent nucleobase that bridges two strands simultaneously. This structural parameter change allows the molecule to achieve high binding affinity through cooperative base-pairing interactions while maintaining the simplicity of a small molecule rather than a complex protein structure
Solution Approach 2:
The connector molecule serves as an intermediary that links two monovalent nucleobases, enabling them to work cooperatively to recognize and bind to mismatched sequences. This intermediary structure facilitates high-affinity binding through simultaneous base-pairing interactions without requiring the complex tertiary structure of proteins
3Stability of the object's composition
If RNA secondary structures are targeted with canonical base-pairing molecules, then binding to stable regions is achieved, but inability to effectively bind to thermodynamically less stable regions occurs
Solution Approach 1:
By segmenting the recognition function into two separate monovalent nucleobases that independently bind to each strand, the divalent nucleobase can target thermodynamically less stable regions where individual base-pairing interactions are weaker. The cooperative binding of both nucleobases compensates for the reduced stability of mismatched or non-canonical regions, enabling targeting across a broader range of RNA structures
Solution Approach 2:
The composite structure of the divalent nucleobase, combining two nucleobase recognition elements with a flexible connector, allows adaptation to various RNA secondary structures including loops, bulges, and junctions. This composite design provides both the stability needed for binding and the versatility to target thermodynamically less stable regions that are crucial for RNA function
Applied Scientific Principles
This section explains which scientific principles are used to turn an abstract innovation direction into a practical engineering solution.
Function Achieved in This Case
Enables the creation of genetic recognition reagents that can target and manipulate nucleic acid sequences, facilitating advanced biological research, diagnostics, and therapeutic applications by enhancing the binding free energy and specificity to RNA structures.
Implementation Method 1
Divalent nucleobases are capable of forming directional hydrogen bonding interactions with two strands of DNA and/or RNA
Data Source
AI summary
Described herein are divalent nucleobases that each binds two nucleic acid strands, matched or mismatched when incorporated into a nucleic acid or nucleic acid analog backbone, such as in a γ-peptide nucleic acid (γPNA). Also provided are genetic recognition reagents comprising one or more of the divalent nucleobases and a nucleic acid or nucleic acid analog backbone, such as a γPNA backbone. Uses for the divalent nucleobases and monomers and genetic recognition reagents containing the divalent nucleobases also are provided.