Abstract
The base excision repair pathway removes small base lesions from DNA and is initiated in humans by one of eleven DNA glycosylases with overlapping substrate specificities. Despite their central role in genome maintenance and transcription, systematic datasets describing small-molecule binders or inhibitors of DNA glycosylases are lacking. Here, through a collaborative effort across Scandinavia, we establish a high-throughput biochemical assay platform for human DNA glycosylases and use it to generate a systematic small-molecule screening dataset. A chemogenomic library was screened against nine glycosylases with assays of sufficient quality for hit identification, yielding primary screening and concentration-response data and identifying multiple previously unreported inhibitors. All primary and processed data, along with detailed assay protocols and metadata, are publicly available to support reuse in chemical biology, further compound optimization, assay development and comparative studies of DNA repair enzymes.