Critical Analysis of Rate Constants and Turnover Frequency in Nucleic Acid-Templated Reactions: Reaching Terminal Velocity.

Nucleic acid-templated reactions have attracted significant attention for nucleic acid sensing and imaging. The level of signal amplification obtained from templated reactions is a function of the template turnover, wherein the template acts as the catalyst. Herein, we report the application of a pyridinium linker that immolates upon photocatalytic reduction with a ruthenium complex to yield the fastest nucleic acid templated reaction reported to date. We show that the templated reaction turnover is limited by the duplex dissociation kinetics beyond probes longer than a 6-mer and proceeded fastest for a 5-mer PNA probe. Using a beacon architecture that masks the catalytic template, we show that this methodology can be used for nucleic acid sensing extending the analyte recognition beyond a 5-mer. The system proceeds with a catalytic efficiency of 105 M-1 s-1 and achieves turnover frequency of >100 h-1.