Supernova: A Deoxyribozyme that Catalyzes a Chemiluminescent Reaction

Abstract

Functional DNAm olecules are useful components in nanotechnology and synthetic biology.T oexpand the toolkit of functional DNAp arts,i nt his study we used artificial evolution to identify ag lowing deoxyribozyme called Supernova. This deoxyribozyme transfers ap hosphate from a1 ,2dioxetane substrate to its 5' hydroxylg roup,w hich triggers ac hemiluminescent reaction and af lash of blue light. An engineered version of Supernova is only catalytically active in the presence of an oligonucleotide complementary to its 3' end, demonstrating that light production can be coupled to ligand binding.W eanticipate that Supernova will be useful in awide variety of applications,i ncluding as as ignaling component in allosterically regulated sensors and in logic gates of molecular computers.

Optimizing the Chemiluminescence of a Light‐Producing Deoxyribozyme

“…Removal of cerium and lead either individually or together had only a small effect on activity (Figures 2A, S2A, and S3A). This was somewhat surprising, since both of these metal ions strongly promote the nonenzymatic dephosphorylation of CDP‐Star at high concentrations [10] . Removal of potassium modestly decreased the rate enhancement of light production of Supernova, while removal of zinc resulted in a complete loss of activity (Figures 2A, S2A, and S3A).…”

“…In addition to providing information about how to maximize light production, our experiments also provide clues about the possible mechanism of the reaction. For example, CDP‐Star titration experiments generate curves that can be fit using the Michaelis‐Menten equation [10] . This indicates that Supernova contains a binding site for CDP‐Star, and suggests that proximity between the nucleophilic 5′ hydroxyl group of Supernova and CDP‐Star could be one mechanism by which Supernova promotes self‐phosphorylation (Figure 8).…”

“…We recently developed a chemiluminescent deoxyribozyme called Supernova, and showed that it can be programmed to produce light only in the presence of a target oligonucleotide [10] . We are also working to create sensors that detect other ligands using Supernova as a starting point.…”

“…Inspired by the many applications of light‐producing reactions, we recently developed a deoxyribozyme that generates light using the triggered 1,2‐dioxetane substrate CDP‐Star (Figure 1). [10] Our deoxyribozyme, called Supernova, transfers the phosphate protecting group from CDP‐Star to its 5′ hydroxyl group, which triggers a decomposition reaction and the production of blue light. We also developed a programmable version of Supernova that only produce light in the presence of specific oligonucleotide sequences [10] .…”

“… [10] Our deoxyribozyme, called Supernova, transfers the phosphate protecting group from CDP‐Star to its 5′ hydroxyl group, which triggers a decomposition reaction and the production of blue light. We also developed a programmable version of Supernova that only produce light in the presence of specific oligonucleotide sequences [10] . This shows that Supernova can be used to detect ligands in solution and raises the possibility that similar sensors could be developed for other ligands.…”