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Dna Synapsis Through Transient Tetramerization Triggers Cleavage By Ecl18ki Restriction Enzyme., Mindaugas Zaremba, Amelia Owsicka, Gintautas Tamulaitis, Giedrius Sasnauskas, Luda S. Shlyakhtenko, Alexander Y. Lushnikov, Yuri L. Lyubchenko, Niels Laurens, Bram Van Den Broek, Gijs J.L. Wuite, Virginijus Siksnys

Journal Articles: Pharmaceutical Sciences

To cut DNA at their target sites, restriction enzymes assemble into different oligomeric structures. The Ecl18kI endonuclease in the crystal is arranged as a tetramer made of two dimers each bound to a DNA copy. However, free in solution Ecl18kI is a dimer. To find out whether the Ecl18kI dimer or tetramer represents the functionally important assembly, we generated mutants aimed at disrupting the putative dimer-dimer interface and analysed the functional properties of Ecl18kI and mutant variants. We show by atomic force microscopy that on two-site DNA, Ecl18kI loops out an intervening DNA fragment and forms a tetramer. Using the …