Nucleic Acids Research Advance Access originally published online on March 30, 2009
Nucleic Acids Research 2009 37(10):3377-3390; doi:10.1093/nar/gkp195
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Nucleic Acids Research, 2009, Vol. 37, No. 10 3377-3390
© 2009 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 |
Type I restriction endonucleases are true catalytic enzymes
Department of Microbiology and Immunology, Department of Biochemistry, Center for Single Molecule Biophysics, The State University of New York at Buffalo, Buffalo, NY 14214, USA
*To whom correspondence should be addressed. Tel: +1 716 829 2599; Fax: +1 716 829 2158; Email: pbianco{at}buffalo.edu
Received January 1, 2009. Revised February 27, 2009. Accepted March 11, 2009.
Type I restriction endonucleases are intriguing, multifunctional complexes that restrict DNA randomly, at sites distant from the target sequence. Restriction at distant sites is facilitated by ATP hydrolysis-dependent, translocation of double-stranded DNA towards the stationary enzyme bound at the recognition sequence. Following restriction, the enzymes are thought to remain associated with the DNA at the target site, hydrolyzing copious amounts of ATP. As a result, for the past 35 years type I restriction endonucleases could only be loosely classified as enzymes since they functioned stoichiometrically relative to DNA. To further understand enzyme mechanism, a detailed analysis of DNA cleavage by the EcoR124I holoenzyme was done. We demonstrate for the first time that type I restriction endonucleases are not stoichiometric but are instead catalytic with respect to DNA. Further, the mechanism involves formation of a dimer of holoenzymes, with each monomer bound to a target sequence and, following cleavage, each dissociates in an intact form to bind and restrict subsequent DNA molecules. Therefore, type I restriction endonucleases, like their type II counterparts, are true enzymes. The conclusion that type I restriction enzymes are catalytic relative to DNA has important implications for the in vivo function of these previously enigmatic enzymes.