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Nucleic Acids Research Advance Access published online on December 17, 2007
Nucleic Acids Research, doi:10.1093/nar/gkm1090
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© 2007 The Author(s)
This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/2.0/uk/) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.

Nucleic Acid Enzymes

Tetrameric restriction enzymes: expansion to the GIY-YIG nuclease family

Giedrius Gasiunas1, Giedrius Sasnauskas1, Gintautas Tamulaitis1, Claus Urbanke2, Dalia Razaniene3 and Virginijus Siksnys1,*

1Institute of Biotechnology, Graiciuno 8, LT-02241 Vilnius, Lithuania, 2Abteilung für Strukturanalyse, Medizinische Hochschule Hannover, Carl Neuberg Strasse 1, D-30632 Hannover, Germany and 3Fermentas UAB, Graiciuno 8, LT-02241 Vilnius, Lithuania

*To whom correspondence should be addressed. Tel: +37 0 5260 2108; Fax: +37 0 5260 2116; Email: siksnys{at}ibt.lt

Received October 11, 2007. Revised November 21, 2007. Accepted November 21, 2007.

The GIY-YIG nuclease domain was originally identified in homing endonucleases and enzymes involved in DNA repair and recombination. Many of the GIY-YIG family enzymes are functional as monomers. We show here that the Cfr42I restriction endonuclease which belongs to the GIY-YIG family and recognizes the symmetric sequence 5'-CCGC/GG-3' (‘/’ indicates the cleavage site) is a tetramer in solution. Moreover, biochemical and kinetic studies provided here demonstrate that the Cfr42I tetramer is catalytically active only upon simultaneous binding of two copies of its recognition sequence. In that respect Cfr42I resembles the homotetrameric Type IIF restriction enzymes that belong to the distinct PD-(E/D)XK nuclease superfamily. Unlike the PD-(E/D)XK enzymes, the GIY-YIG nuclease Cfr42I accommodates an extremely wide selection of metal-ion cofactors, including Mg2+, Mn2+, Co2+, Zn2+, Ni2+, Cu2+ and Ca2+. To our knowledge, Cfr42I is the first tetrameric GIY-YIG family enzyme. Similar structural arrangement and phenotypes displayed by restriction enzymes of the PD-(E/D)XK and GIY-YIG nuclease families point to the functional significance of tetramerization.

The authors wish it to be known that, in their opinion, the first two authors should be regarded as joint First Authors


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