Sortase-Mediated Nucleic Acid Conjugation to Cell Surfaces
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Solution Overview
Problem
Existing technologies have not explored the potential of nucleic acids or their derivatives as substrates for sortase-mediated reactions, limiting the applications of sortase in bioconjugation and cellular interactions.
Innovation Solution
The discovery that nucleic acids, such as DNA and RNA, or their derivatives can serve as substrates for sortase, enabling their stable anchoring to cell surfaces through a sortase-mediated reaction, which was previously unknown.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Adaptability or versatility
If sortase is used for bioconjugation with traditional peptide substrates, then protein attachment to cell surfaces is achieved, but the application scope is limited to proteins and peptides only
Solution Approach 1:
The sortase enzyme is engineered to perform multiple functions: it maintains its original ability to process peptide substrates with LPXTG motifs while simultaneously acquiring the capability to process nucleic acid substrates. This multi-functionality allows a single enzyme system to mediate conjugation of diverse molecules including proteins, peptides, DNA, and RNA to cell surfaces, thereby expanding application scope without sacrificing reaction reliability
Solution Approach 2:
The invention modifies the substrate recognition parameters of sortase by introducing engineered variants with altered specificity. These variants can recognize different motifs beyond the canonical LPXTG peptide sequence, including nucleic acid structures. By changing the recognition parameters while maintaining catalytic function, the enzyme achieves broader substrate compatibility while preserving reliable and specific conjugation reactions
2Reliability
If nucleic acids are conjugated to cells for therapeutic applications, then targeted drug delivery and cellular labeling are enabled, but the stability of nucleic acids on cell surfaces is challenging
Solution Approach 1:
The sortase-mediated conjugation system performs preliminary anchoring of nucleic acids to cell surfaces through specific enzymatic recognition and covalent bond formation. This preliminary action creates stable, site-specific attachment points that prevent nucleic acid detachment, thereby ensuring reliable anchoring stability while maintaining a relatively simple and controlled conjugation process through enzymatic specificity
Solution Approach 2:
The sortase enzyme acts as an intermediary that facilitates the conjugation between nucleic acids and cell surfaces. It mediates the reaction by recognizing specific motifs on the nucleic acid substrates and catalyzing their covalent attachment to cell surface proteins or structures. This intermediary role ensures stable anchoring while simplifying the overall process through enzyme-catalyzed specificity, reducing the need for complex chemical modifications
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
This novel reaction allows for the efficient conjugation of nucleic acids to cells, facilitating their internalization and potential therapeutic applications, such as targeted drug delivery and cellular labeling.
Implementation Method 1
peptides with an LPXTG motif were reported to be recognized by SrtA and were covalently anchored to an NH2-GGG peptide on the cell wall through a transpeptidation reaction
Implementation Method 2
Sortase (Srt), e.g., Sortase A (SrtA)... is a group of transpeptidases that mediate attaching peptides to bacteria cell walls
Data Source
AI summary
The present disclosure provides a conjugate of a nucleic acid or derivative thereof and a sortase. The present disclosure also provides a conjugate of a nucleic acid or derivative thereof and a cell, and a method of preparing such a conjugate mediated by a sortase. The present disclosure further provides a method of delivering a nucleic acid or derivative thereof to a cell, mediated by a sortase.