Abasic site ring opening and DNA-protein crosslink reversal by the SRAP protein YedK

Apuirinic/apyrimidinic (AP, or abasic) sites in DNA are one of the most common forms of DNA damage. AP sites are reactive and form crosslinks to both proteins and DNA, are prone to strand breakage, and inhibit DNA replication and transcription. The protein HMCES protects cells from strand breaks, inhibits mutagenic translesion synthesis, and participates in repair of interstrand DNA crosslinks derived from AP sites by forming a stable thiazolidine DNA-protein crosslink (DPC) to AP sites in single-stranded DNA (ssDNA). Despite the importance of HMCES to genome maintenance and the evolutionary conservation of its catalytic SRAP (SOS Response Associated Peptidase) domain, the enzymatic mechanisms of DPC formation and resolution are unknown. Using the bacterial homolog YedK, we show that the SRAP domain catalyzes conversion of the AP site to its reactive, ring-opened aldehyde form, and provide structural evidence for the Schiff base intermediate that forms prior to the more stable thiazolidine. We also report two new activities, whereby SRAP reacts with polyunsaturated aldehydes at DNA 3-ends generated by bifunctional DNA glycosylases and catalyzes direct reversal of the DPC to regenerate the AP site, which provide insight into possible mechanisms by which HMCES DPCs are resolved in cells.