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ATP-dependent type I topoisomerases from extremely thermophilic archaebacteria--reverse gyrases--drive positive supercoiling of DNA. We showed that reverse gyrase from Desulfurococcus amylolyticus breaks the DNA at specific sites and covalently binds to the 5' end. In 30 out of 31 sites located in pBR322 DNA fragments, cleavage occurs at the sequence 5'---CNNN/---(N is any base). The same rule was previously shown to hold for single-stranded DNA breakage by eubacterial topoisomerases I. The relative cleavage frequencies at different sites depend on Mg2+ and temperature. We discuss the possible physiological and mechanistic role of the above specificity of the bacterial topoisomerases I.

Archaebacterial reverse gyrase cleavage-site specificity is similar to that of eubacterial DNA topoisomerases I