Nucleic Acids Research Advance Access published online on September 29, 2006
Nucleic Acids Research, doi:10.1093/nar/gkl489
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© 2006 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.
Molecular Biology |
Structural and functional analysis of the MutS C-terminal tetramerization domain
Institut für Biochemie, Justus-Liebig-Universität Heinrich-Buff-Ring 58, D-35392 Giessen, Germany 1 Medizinische Hochschule, Strukturanalyse Carl Neuberg Strasse 1, D-30625 Hannover, Germany
*To whom correspondence should be addressed: Tel: +49 641 99 35407; Fax: +49 641 99 35409; Email: friedhoff{at}chemie.bio.uni-giessen.de
Received May 22, 2006. Revised June 27, 2006. Accepted June 27, 2006.
The Escherichia coli DNA mismatch repair (MMR) protein MutS is essential for the correction of DNA replication errors. In vitro, MutS exists in a dimer/tetramer equilibrium that is converted into a monomer/dimer equilibrium upon deletion of the C-terminal 53 amino acids. In vivo and in vitro data have shown that this C-terminal domain (CTD, residues 801–853) is critical for tetramerization and the function of MutS in MMR and anti-recombination. We report the expression, purification and analysis of the E.coli MutS-CTD. Secondary structure prediction and circular dichroism suggest that the CTD is folded, with an 
-helical content of 30%. Based on sedimentation equilibrium and velocity analyses, MutS-CTD forms a tetramer of asymmetric shape. A single point mutation (D835R) abolishes tetramerization but not dimerization of both MutS-CTD and full-length MutS. Interestingly, the in vivo and in vitro MMR activity of MutSCF/D835R is diminished to a similar extent as a truncated MutS variant (MutS800, residues 1–800), which lacks the CTD. Moreover, the dimer-forming MutSCF/D835R has comparable DNA binding affinity with the tetramer-forming MutS, but is impaired in mismatch-dependent activation of MutH. Our data support the hypothesis that tetramerization of MutS is important but not essential for MutS function in MMR.
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