Helical-helicase 2 domains interactions in HsdR subunit of EcoR124I restriction-modification complex


V. Bialevich1,2, M. Weiserova3, R. Ettrich1,2 and E. Csefalvay1,2


1Institute of Nanobiology and Structural Biology of ISBE, AS CR, 37333 Nove Hrady, Czech Republic

2University of South Bohemia in České Budějovice, 37333 Nove Hrady, Czech Republic

3Institute of Microbiology, AS CR, Vídeňská 1083, 142 20, Praha 4


Procaryotic type I Restriction-Modification systems effectively recognize and destroy phage DNA by cooperative recruitment of endonuclease, ATPase and DNA translocase and protect the host genome from being self-restricted by DNA methyltransferase action [1].

Type I R-M systems are multi-subunit enzymes. The complex is composed of two HsdR (Restriction and translocation) subunits, two HsdM (Methylation) subunits, and one HsdS (Specifity) subunit. HsdR subunit is the biggest part of the complex and responsible for DNA cleavage. Additionally it acts as an ATP-dependent DNA translocase [1]. HsdR is organized into four approximately globular structural domains in nearly square-planar arrangement: the N-terminal endonuclease domain, the recA-like helicase domains 1 and 2 and the C-terminal helical domain. The near-planar arrangement of globular domains creates prominent grooves between each domain pair. The two helicase-like domains form a canonical helicase cleft in which double-stranded B-form DNA can be accommodated without steric clash. A positively charged surface groove proceeds from the helicase cleft and continues between the helical and endonuclease domains where it passes over the cleavage site recessed slightly from the surface [2]. The C-terminal helical domain resembles the fold of HsdM and has strong interactions with helicase 2 domain. During translocation large-scale rotation of helicase domain 2 relative to the helicase domain 1 is expected, which would significantly alter the helical-helicase domain interface. Salt-bridges or hydrogen bonds over this interface are the major energetic contribution to the binding energy between both domain, and must play a key role in accommodating the different rotational stages of helicase domain 2 during the translocation cycle. We map residues that are essential for helical-helicase 2 domains interactions by using a combination of site-directed mutagensis and in vivo and in vitro activity assays and put the results into the broader context of DNA translocation and following restriction.


1.       Murray, N. E. Microbiology and Molecular Biology Reviews, June 2000, Vol. 64, No. 2, p. 412–434.

2.       Lapkouski M., Panjikar S., Janscak P., Kuta Smatanova I., Carey J., Ettrich R., Csefalvay E. Nat. Struct. & Mol.Biol, 2009, 16, 94.


We gratefully acknowledge support from the Ministry of Education, Youth and Sports of the Czech Republic (MSM6007665808, LC06010), the Academy of Sciences of the Czech Republic (AVOZ60870520), the Grant Agency of the Czech Republic (Nos. P207/10/1934), and joint Czech - US National Science Foundation International Research Cooperation (ME09016 and INT03-09049), Additionally, V.B. was supported by the University of South Bohemia, grant GAJU 170/2010/P.