Design, synthesis, and assembly of sequence-defined vinyl sulfonamide click nucleic acids (VS-CNAs)
Date
2019
Authors
Journal Title
Journal ISSN
Volume Title
Publisher
University of Delaware
Abstract
DNA is a vast source of inspiration in the design of highly programmable materials with specific functions and assembly properties. Through sequence-specific hydrogen bonding interactions, DNA can encode all genetic information about a living organism. Despite DNA’s ability to store information and direct the assembly of intricate nanostructures, its uses in engineering applications are limited owing to both scale and cost. Additionally, the phosphate ribose-based backbone structure of DNA is lacks chemical robustness and is readily degraded by the enzymes. Inspired by the hydrogen bonding interactions found in natural DNA, biomimetic polymers with pendent nucleobases have been developed to design and create programmable materials. However, many of these polymeric materials lack the ability to impart sequence specificity, limiting the degree of assembly. Click nucleic acids (CNAs) use highly efficient thiol-based click conjugation reactions to enable the synthesis of nucleobase sequences with high fidelity. Unfortunately, the thiol-acrylamide and thiol-halogen conjugation reactions used in current CNA monomers exhibit poor kinetics and produce considerable side products. Additionally, to date only thymine and adenine monomers have been synthesized limiting the range of sequences that can be incorporated into CNA polymers. ☐ An improved thiol-Michael acceptor is proposed and incorporated into CNA monomer structures to improve the sequence conjugation strategy. Model kinetic studies showed vinyl sulfonamides exhibited two orders of magnitude higher reactivity than acrylamides without sacrificing bond stability, as opposed to acrylates and maleimides, making them a suitable replacement. The effectiveness of the vinyl sulfonamide was demonstrated in the application of on-resin macrocyclization of a model RGDS peptide. Successful macrocyclization was achieved with over 95% efficiency confirming the vinyl sulfonamide as a highly efficient thiol-Michael acceptor. Vinyl sulfonamide functional handles were readily incorporated onto all four nucleobase containing monomers in good yields with minimal chromatographic purification steps. To achieve sequence control, a solution-phase approach was adopted using thiol functional poly(ethylene glycol) (PEG) as a polymer support. Iterative conjugation and deprotection steps resulted in near quantitative yields for both the thiol-Michael reaction and removal of the protecting group. Additionally, building CNAs from the PEG polymer support enables a simple and scalable purification method. Furthermore, the solution-phase approach was applied to thiol functional multiarm PEG macromers, to create PEG-CNA sequences that can assemble with complementary sequences into a dynamically responsive hydrogel. Through strand displacement, these CNA hydrogels degraded in the presence of complementary linear PEG-CNAs. Successful design of sequence-specific CNAs with all four nucleobases further advances their applicability as a DNA mimetic in the design of sequence-directed assemblies. ☐ The evaluation of the CNA monomer structure lead to an unexpected acid-catalyzed thiol-ene-type reaction that proceeds via a cationic mechanistic pathway. Using model studies, various thiol and vinyl substrates were evaluated to determine the scope of the acid-catalyzed thiol-vinyl reaction. The kinetics were fit to a Michaelis-Menten model as the reaction demonstrated saturation kinetics with rapid equilibrium of multiple intermediate products followed by a rate determining sulfonium dissociation step. Orthogonality of the cationic-mediate vinyl-thiol reaction with the radical mediated thiol-ene and anionic mediated thiol-Michael reactions, enabled a demonstration of three sequential thiol-X reactions in a one-pot scheme. Furthermore, photoinitiation could be achieved using a photochromic photoacid that produces an acidic proton upon isomerization under 405 nm light. The expansion of the thiol-X toolbox with the addition of the cationic-mediated thiol-vinyl reaction advances the development of next generation functional materials and thiol conjugation strategies.
Description
Keywords
Dynamic, Nucleic acid, Nucleobase, Peptide, Polymer, Sequence control