Evolution of sequence-defined highly functionalized nucleic acid polymers

Abstract

The evolution of sequence-defined synthetic polymers made of building blocks beyond those compatible with polymerase enzymes or the ribosome has the potential to generate new classes of receptors, catalysts, and materials. Here we describe a ligase-mediated DNA-templated polymerization system and in vitro selection to evolve highly functionalized nucleic acid polymers (HFNAPs) made from 32 building blocks containing eight chemically diverse side-chains on a DNA backbone. Through iterated cycles of polymer translation, selection, and reverse translation, we discovered HFNAPs that bind PCSK9 and IL-6, two protein targets implicated in human diseases. Mutation and reselection of an active PCSK9-binding polymer yielded evolved polymers with high affinity (KD = 3 nM). This evolved polymer potently inhibited binding between PCSK9 and the LDL receptor. Structure-activity relationship studies revealed that specific side-chains at defined positions in the polymers are required for binding to their respective targets. Our findings expand the chemical space of evolvable polymers to include densely functionalized nucleic acids with diverse, researcher-defined chemical repertoires.

DNA-Programmed Chemical Synthesis of Polymers and Inorganic Nanomaterials

“…Liu and other researchers also attempted to mimic the principle of the natural gene translation process to achieve chemical synthesis of sequence-controlled polymers based on DNA templates [29][30][31]. They reported the efficient and sequencespecific polymerization of non-functional [32] and side-chain-functionalized peptide nucleic acid aldehydes [33] with DNA sequence templates, combined with an in vitro translation, functional screening and amplification system [34].…”

Beyond DNA and RNA: The Expanding Toolbox of Synthetic Genetics

“…Again, as described above, this approach may also benefit from the restricted rotation of the resulting 1,2,3-triazole ring. In a different approach, "synthetic translation" has the potential to offer an even wider range of substituent chemistries: DNA triplets bearing one of eight different side chains can be aligned on DNA templates and ligated (by T4 DNA ligase) to prepare heavily modified DNA, which was recently used to select aptamers against IL-6 and PCSK9 (Chen et al 2018).…”

Nucleic Acid‐Barcoding Technologies: Converting DNA Sequencing into a Broad‐Spectrum Molecular Counter

“…[35] Other notable approaches for DNA-templated synthesis include the YoctoReactor [36] and densely functionalized nucleic-acid polymers. [37] Much like the Liu approach described above,the YoctoReactor exploits DNAhybridization to direct chemical reactions through modulating effective molarity.T his method is unique in that the DNA-encoded building blocks are programmed to position their respective reactive chemical moieties at the center of three-way DNA junctions to achieve an unprecedented reaction volume (1 yoctoliter (10 À24 L)). [36] Densely functionalized nucleic-acid polymers are DNApolymers in which every third nucleotide contains af unctionalized base moiety.T oa ssemble such libraries in as ingle-pot, functionalized trinucleotides (or codons) are prepared and hybridized with alibrary of singlestranded DNAt emplates that present unique codon reading frames.After synthetic-codon annealing, DNAl igase is used to ligate the codons and create the functionalized nucleic-acid polymer.Excellent reviews further describing these and other DNA-encoded small molecule library technologies and their success in drug discovery can be found elsewhere.…”