# Amphiphilic Oligonucleotide Derivatives as a Tool to Study DNA Repair Proteins

## TL;DR

This paper explores how modified DNA molecules with lipophilic groups interact with DNA repair proteins, offering new tools for studying these proteins in cells.

## Contribution

The study introduces amphiphilic DNA duplexes with lipophilic substituents as novel tools for analyzing DNA repair proteins like PARPs.

## Key findings

- LS-DNA binds more efficiently to PARP1, PARP2, PARP3, and DNA polymerase β compared to regular DNA.
- Photo-LS-DNAs enable affinity labeling of PARPs and tracking of Ku antigen in different cell types.
- PARP1–PARP3 can slowly excise 5′ dRP groups at DNA breaks, and LS-DNAs activate PARP1 and PARP2 for autoPARylation.

## Abstract

Modified oligonucleotides (oligos) are widely used as convenient tools in many scientific fields, including biomedical applications and therapies. In particular, oligos with lipophilic groups attached to the backbone ensure penetration of the cell membrane without the need for transfection. This study examines the interaction between amphiphilic DNA duplexes, in which one of the chains contains a lipophilic substituent, and several DNA repair proteins, particularly DNA-damage-dependent PARPs, using various biochemical approaches. DNA with a lipophilic substituent (LS-DNA) demonstrates more efficient binding with DNA damage activated poly(AD-ribose) polymerases 1-3 (PARP1, PARP2, PARP3) and DNA polymerase β. Chemically reactive LS-DNA derivatives containing a photoactivatable nucleotide (photo-LS-DNAs) or a 5′ deoxyribose phosphate (dRP) group in the vicinity of double-strand breaks (DSBs) are used for the affinity labelling of PARPs and other proteins in several whole-cell extracts of human cells. In particular, photo-LS-DNAs are used to track the level of Ku antigen in the extracts of neuron-like differentiated SH-SY5Y, undifferentiated SH-SY5Y, and olfactory epithelial cells. In vitro, PARP1–PARP3 are shown to be able to slowly excise the 5′ dRP group at DSBs. LS-DNAs can activate PARP1 and PARP2 for autoPARylation, albeit less effectively than regular DNA duplexes.