Antibody Activation using DNA‐Based Logic Gates

Abstract

Oligonucleotide‐based molecular circuits offer the exciting possibility to introduce autonomous signal processing in biomedicine, synthetic biology, and molecular diagnostics. Here we introduce bivalent peptide–DNA conjugates as generic, noncovalent, and easily applicable molecular locks that allow the control of antibody activity using toehold‐mediated strand displacement reactions. Employing yeast as a cellular model system, reversible control of antibody targeting is demonstrated with low nM concentrations of peptide–DNA locks and oligonucleotide displacer strands. Introduction of two different toehold strands on the peptide–DNA lock allowed signal integration of two different inputs, yielding logic OR‐ and AND‐gates. The range of molecular inputs could be further extended to protein‐based triggers by using protein‐binding aptamers.

Antibody-controlled actuation of DNA-based molecular circuits

“…First, the product of the exchange reaction BI can form a bivalent interaction with its target antibody, thus making the displacement of O by I thermodynamically more favourable. We recently showed that bivalent peptide-dsDNA ligands form very tight 1:1 complexes with their target antibody, showing a 500-fold increase in affinity compared to the monovalent peptide-antibody interaction 33 34 . Second, the colocalization of the reactants increases their effective concentration, hence enhancing the rate of the exchange reaction.…”

“…To ensure minimal background in combination with fast initiation of the ATSE reaction, we optimized the design by systematically increasing the length of the toehold T from 0 to 6 nucleotides, hence gradually destabilizing the initial state. The length of the strands was chosen such that the bivalent peptide-dsDNA product BI contained 24 basepairs, a length that we have previously shown to be optimal for bivalent binding to the target antibody 33 34 . In the absence of input antibody no increase in fluorescence was observed up to a toehold length of 3 nucleotides ( Fig.…”

“…Both thermodynamic and kinetic effects contribute to the remarkable efficiency of ATSE. First, the bivalent peptide-dsDNA product of the ATSE reaction forms a highly stable 1:1 cyclic complex with its bivalent target antibody, thus making the displacement reaction thermodynamically favourable 33 34 . Second, colocalization of the peptide-functionalized oligonucleotides on the two antigen binding domains increases their effective concentration, hence enhancing the rate of the exchange reaction.…”

Protein–Protein Communication Mediated by an Antibody‐Responsive DNA Nanodevice**

“…25,26 Synthetic oligonucleotides can also be conjugated with other recognition elements responding to a wide range of proteins and biomolecules, further broadening the potential interface between the world of synthetic nucleic acids and proteins. Recently, we and others have, for example, employed antigenconjugated synthetic DNA strands to allow programmable interactions with specific antibodies [27][28][29][30][31] that control the assembly and disassembly of DNA-based molecular structures. 32 In the above-described examples, the communication is limited to protein-to-DNA interactions in which a specific protein (e.g., a transcription factor, an enzyme, or an antibody) triggers a functional event in a structure built of DNA.…”

Programmable cell-free transcriptional switches for antibodies detection

“…Synthetic nucleic acid strands can be used as molecular scaffolds to append different recognition elements that allow the design of nucleic acid probes responsive to a wide range of targets. [15][16][17][18][19] In a demonstration of such potentiality, we and others have recently employed antigen-conjugated nucleic acids rationally designed to respond to clinically-relevant antibodies. [19][20][21][22][23] Motivated by the above considerations, we demonstrate here a cell-free diagnostic platform for the detection of specific antibodies in blood serum based on the use of antibody-responsive nucleic acid transcriptional switches.…”