Template‐Directed Ligation: Towards the Synthesis of Sequence Specific Polymers

Abstract

The structure of the DNA double helix first highlighted the simple elegance of the most sophisticated template reactions known, template directed polymerization. Through these reactions the genomic information is accurately replicated in DNA, transcribed into RNA, and translated into protein. Over 40 years ago, Crick combined these reactions into the "Central Dogma" of biology ( Figure 5-l), a formulation suggesting that all living things follow this basic chemical plan [ 11. Today, even with the incredible biological diversity known to exist on Earth, no violations of this chemical scheme are known. The nucleic acids store the genetic blueprint, and the co-catalytic proteins function cooperatively with this molecular template to express the encoded information.Over the last four decades, synthetic non-enzymatic oligonucleotide ligations have been explored as potential models for the evolution of replicating systems on the early Earth [2-111. These studies have explored ligation reactions that occur on a template without protein/enzymatic catalysis. Such systems have both provided mechanisms for the early evolution of biopolymer catalysts and placed limits on template directed ligation. More recently, the lure of antisense molecules capable of disrupting specific gene expression has resulted in the exploration of backbone-modified nucleic acids [ 12-33]. n

Efficient and Sequence-Specific DNA-Templated Polymerization of Peptide Nucleic Acid Aldehydes

“…The highly efficient and sequence-specific nucleic acid-templated polymerization of proteins and nucleic acids is a fundamental feature of living systems that enables biopolymers to evolve . Over the past few decades, several efforts to adapt templated polymerization to synthetic systems have been reported . These systems typically use oligonucleotides (or analogues) as a template for coupling DNA- or RNA-based monomers.…”

Digital and Analog Chemical Evolution

“…Like the phosphodiester, imine condensation releases water and is thermodynamically disfavored in aqueous environments, but the lower activation barrier has been exploited in the creation of one of the earliest examples of a synthetic dynamic chemical network (Figure ). ,− The network responds to a complementary template, and hydride reagents are used to chemospecifically trap only the template-bound imine. The binding affinity of the resulting secondary amine-linked DNA oligomers to the DNA template ( K 5 ) is 10 6 -fold lower than the ternary substrate/template complex ( K 3 ), leading to isothermal dissociation of the product strand from the template and catalytic turnover (Figure ). ,,, When sequence-complementary templates are introduced to this dynamic network at a 1% molar equivalent relative to substrate, efficient product turnover reveals a long-sought strategy to overcome the critical product inhibition limitation of digital molecular information storage in nucleic acids …”

Step-Growth Control in Template-Directed Polymerization

“…This process has been simplified by combining reversible imine condensation on a template with kinetic reductive amination for controlling polymer growth. [13][14][15][16] Specifically, thymidine carrying a 5'-amine and a 3'-acetaldehyde, 5'-H 2 N-dT-3'-CH 2 CHO, T 1 , provides the polymerization monomer ( Figure 1). While T 1 can form intramolecular imines, no reaction is detected under mild reducing conditions in aqueous environments.…”

“…Such a progressive reduction in the number of sites will impact the imine concentration and the reduction rates. Maybe more importantly, the amine backbone oligomers are known to associate more weakly to a DNA template than the imine, 13,16 further impacting the concentration of template-bound product early in oligomerization. Accordingly, we have constructed a model for the reaction using nearest-neighbor thermodynamic parameters for DNA Watson-Crick base pairs 17 to estimate equilibrium constants, and Table 2 lists the parameters used in the calculations.…”