Expanded Substrate Scope of DNA Polymerase θ and DNA Polymerase β: Lyase Activity on 5′-Overhangs and Clustered Lesions

• Published in: Biochemistry (Easton) 2018-10, Vol.57 (42), p.6119-6127
• Main Authors: Laverty, Daniel J, Greenberg, Marc M
• Format: Article
• Language: English
• Subjects: DNA Damage; DNA Polymerase beta - chemistry; DNA Polymerase beta - metabolism; DNA Polymerase theta; DNA Repair; DNA Replication; DNA-directed DNA polymerase; DNA-Directed DNA Polymerase - chemistry; DNA-Directed DNA Polymerase - metabolism; enzyme activity; Humans; Phosphorus-Oxygen Lyases - chemistry; Phosphorus-Oxygen Lyases - metabolism; Substrate Specificity
• ISSN: 0006-2960; 1520-4995; 1520-4995
• DOI: 10.1021/acs.biochem.8b00911

DNA polymerase θ (Pol θ) is a multifunctional enzyme with double-strand break (DSB) repair, translesion synthesis, and lyase activities. Pol θ lyase activity on ternary substrates containing a 5′-dRP that are produced during base excision repair of abasic sites (AP) is weak compared to that of DNA polymerase β (Pol β), a polymerase integrally involved in base excision repair. This led us to explore whether Pol θ utilizes its lyase activity to remove 5′-dRP and incise abasic sites from alternative substrates that might be produced during DNA damage and repair. We found that Pol θ exhibited lyase activity on abasic lesions near DSB termini and on clustered lesions. To calibrate the Pol θ activity, Pol β reactivity was examined with the same substrates. Pol β excised 5′-dRP from within a 5′-overhang 80 times faster than did Pol θ. Pol θ and Pol β also incised AP within clustered lesions but showed opposite preferences with respect to the polarity of the lesions. AP lesions in 5′-overhangs were typically excised by Pol β 35–50 times faster than those in a duplex substrate but 15–20-fold more slowly than 5′-dRP in a ternary complex. This is the first report of Pol θ exhibiting lyase activity within an unincised strand. These results suggest that bifunctional polymerases may exhibit lyase activity on a greater variety of substrates than previously recognized. A role in DSB repair could potentially be beneficial, while the aberrant activity exhibited on clustered lesions may be deleterious because of their conversion to DSBs.