Interdomain interaction in the motor subunit of the type I restriction-modification system EcoR124I and their functional relevance

Sinha D^1,2, Bialevich V^1,2, Shamayeva K^1,2, Guzanova A³, Csefalvay E¹, Reha D¹, Weiserova M³, Ettrich R^1,2

¹Institute of Nanobiology and Structural Biology, Global Change Research Center, Academy of Sciences of the Czech Republic, Zamek 136, CZ-373 33 Nove Hrady, Czech Republic

²Faculty of Sciences, University of South Bohemia in Ceske Budejovice, Zamek 136, CZ-373 33 Nove Hrady, Czech Republic

³Institute of Microbiology, Academy of Sciences of the Czech Republic, Vídeňská 1083, 14220 Praha 4, Czech Republic

EcoR124I is a type I restriction-modification (R-M) enzyme and as such forms multifunctional heteromeric complexes with DNA cleavage and ATP-dependent DNA translocation activities located on motor subunit HsdR. When unmethylated invading DNA is recognized by the complex, two HsdR endonuclease/motor subunits start to translocate dsDNA without strand separation activity up to thousands base pairs towards the stationary enzyme while consuming ~1 molecule of ATP per base pair advanced. Finally, after translocation is blocked the HsdR subunits cleave the dsDNA nonspecifically far from recognition site. The crystal structure of the motor subunit R determined by our group in 2009 [1] revealed the four domains within the subunit in a square planer arrangement. Computational modeling including molecular dynamics in combination with crystallography, point mutations, in vivo and in vitro assays reveal how interactions between these four domains contribute to ATP-dependent DNA translocation, DNA cleavage or interdomain communication between the translocase and endonuclease activities [2,3,4,5].

1.         Mikalai Lapkouski, et al.[2009]Structure of the motor subunit of type I restriction-modification complex EcoR124I.,Nat Struct Mol Biol16:1.94-95Jan

2.         D Sinha, et al.[2014]Interdomain communication in the endonuclease/motor subunit of Type I restriction-modification enzyme EcoR124I,Journal of Molecular Modeling20 (7):2334

3.         Eva Csefalvay, et al.[2015]Functional coupling of duplex translocation to DNA cleavage in a type I restriction enzyme PLOS One accepted with minor revisions 21.1.2015.

4.         Bialevich V, et al.The role of motif III and its extended region in positioning the two helicase domains in the motor subunit of the restriction-modification system EcoR124I [manuscript in preparation]

5.         Sinha D, et al.The helical domain of the motor subunit of EcoR124I participates in ATPase activity and dsDNA translocation[manuscript in preparation]