Year: 2025
Type: article
Abstract: This study establishes a real-time fluorescence microscopy platform for visualizing ligand binding dynamics to His-tagged proteins bound to Ni-NTA agarose beads. By preserving solution-phase kinetics while enabling sub-minute temporal resolution in physiological buffers, the methodology overcomes critical limitations of surface-based techniques and gel electrophoretic methods. We applied this platform to investigate inhibitor action within a nucleosomal system, a more physiologically relevant context than free DNA. Through studies of PARP2-nucleosome interactions modulated by clinical inhibitors (talazoparib, veliparib) and by reaction of poly(ADP-ribosyl)ation in the presence of NAD+, we demonstrate direct spatial and temporal resolution of chromatin-protein dynamics. The platform’s virtually unlimited buffer compatibility, real-time monitoring capabilities, and elimination of covalent immobilization artifacts provide transformative insights into drug mechanisms and chromatin engagement processes.
Language: Russian
Subfield: Oncology
Field: Medicine
Domain: Health Sciences
Open Access status: hybrid
Funder: Russian Science Foundation
Awards: G3126765955, G5378482899
OpenAlex ID: W4416308950