Single-site DNA cleavage by Type III restriction endonuclease requires a site-bound enzyme and a trans-acting enzyme that are ATPase-activated

Nucleic Acids Research
Ishtiyaq AhmadKayarat Saikrishnan

Abstract

Endonucleolytic cleavage of DNA by Type III restriction-modification (RM) enzymes requires long-range communication between at least two recognition sites in inverted orientation. This results in convergence of two nuclease domains, one each from the enzymes loaded at the recognition sites with one still bound to the site. The nucleases catalyze scission of the single-strands leading to double-strand DNA break. An obscure feature of the Type III RM enzymes EcoP1I and EcoP15I is their ability to cleave DNA having a single recognition site under certain conditions. Here we demonstrate that single-site cleavage is the result of cooperation between an enzyme bound to the recognition site in cis and one in trans. DNA cleavage is catalyzed by converging nucleases that are activated by hydrolysis-competent ATPase in presence of their respective DNA substrates. Furthermore, a single activated nuclease cannot nick a strand on its own, and requires the partner. Based on the commonalities in the features of single-site and two-site cleavage derived from this study, we propose that their mechanism is similar. Furthermore, the products of two-site cleavage can act as substrates and activators of single-site cleavage. The difference in the t...Continue Reading

References

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Citations

Jul 13, 2019·Nature Communications·Neha NirwanKayarat Saikrishnan
Jul 12, 2020·Journal of Structural Biology·Christopher J HosfordJoshua S Chappie
Apr 12, 2021·Biochemistry. Biokhimii︠a︡·Artem B IsaevKonstantin V Severinov

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