Local Chromatin Organization and Net Charge Modulate DNA Glycosylase Efficiency in Live Cells

Diffusion governs molecular encounter rates and underpins the efficiency of chemical reactions, including those catalyzed by enzymes in crowded cellular environments. In the context of DNA repair, how local nuclear organization modulates enzymatic search efficiency remains poorly understood. Here, we apply spatially resolved single-molecule diffusivity mapping to investigate the dynamics of human 8-oxoguanine DNA glycosylase (hOGG1) in live cell nuclei. We identify three key determinants of enzymatic performance: chromatin compactness, the balance between one-dimensional sliding and three-dimensional hopping, and the 3D diffusion rate during hopping. Approximately 50% of the search process occurs via sliding along chromatin, and a small-molecule activator enhances repair efficiency by inducing chromatin condensation. Guided by theoretical modeling, we further show that reducing the enzyme’s net positive charge increases hopping mobility and improves repair efficiency. These findings demonstrate how chromatin architecture and electrostatic properties govern enzyme dynamics in vivo, offering a general strategy for tuning intracellular reactivity through physicochemical design.