# The Effect of 8,5′-Cyclo 2′-deoxyadenosine on the Activity of 10-23 DNAzyme: Experimental and Theoretical Study

## TL;DR

This study examines how a modified nucleoside affects the activity of a DNAzyme in cleaving RNA, combining experimental and theoretical approaches.

## Contribution

The novel contribution is the investigation of how substituting a specific nucleoside impacts DNAzyme activity and flexibility in physiological conditions.

## Key findings

- Substituting 2′-deoxyadenosine with cdA in the catalytic loop decreased DNAzyme activity.
- The inhibitory effect of RcdA5 may result from increased rigidity in the catalytic loop.
- Theoretical studies showed structural changes in phosphate group distances affecting activity.

## Abstract

The in vivo effectiveness of DNAzymes 10-23 (Dz10-23) is limited due to the low concentration of divalent cations. Modifications of the catalytic loop are being sought to increase the activity of Dz10-23 in physiological conditions. We investigated the effect of 5′S or 5′R 5′,8-cyclo-2′deoxyadenosine (cdA) on the activity of Dz10-23. The activity of Dz10-23 was measured in a cleavage assay using radiolabeled RNA. The Density Functional Tight Binding methodology with the self-consistent redistribution of Mulliken charge modification was used to explain different activities of DNAzymes. The substitution of 2′-deoxyadenosine with cdA in the catalytic loop decreased the activity of DNAzymes. Inhibition was dependent on the position of cdA and its absolute configuration. The order of activity of DNAzymes was as follows: wt-Dz > ScdA5-Dz ≈ RcdA15-Dz ≈ ScdA15-Dz > RcdA5-Dz. Theoretical studies revealed that the distance between phosphate groups at position 5 in RcdA5-Dz was significantly increased compared to wt-Dz, while the distance between O4 of dT4 and nonbonding oxygen of PO2 attached to 3′O of dG2 was much shorter. The strong inhibitory effect of RcdA5 may result from hampering the flexibility of the catalytic loop (increased rigidity), which is required for the proper positioning of Me2+ and optimal activity.