Recording Binding Information Directly into DNA-Encoded Libraries Using Terminal Deoxynucleotidyl Transferase.

ABSTRACT: Terminal deoxynucleotidyl transferase (TdT) is an unusual DNA polymerase that adds untemplated dNTPs to 3'-ends of DNA. If a target protein is expressed as a TdT fusion and incubated with a DNA-encoded library (DEL) in the presence of dATP, the binders of the target induce proximity between TdT and the DNA, promoting the synthesis of a poly-adenine (polyA) tail. The polyA tail length is proportional to the binding affinity, effectively serving as a stable molecular record of binding events. The polyA tail is also a convenient handle to enrich binders with magnetic poly(dT)₂₅ beads before sequencing. In a benchmarking system, we show that ligands spanning nanomolar to double-digit micromolar binding can be cleanly identified by TdT extension, whereas only the tightest binding lig

SUBMITTER: Schneider LA 

PROVIDER: S-EPMC10540198 | biostudies-literature | 2023 Sep

REPOSITORIES: biostudies-literature