Regulatory mutations in Sin recombinase support a structure-based model of the synaptosome

Rowland, S.J., Boocock, M.R., MacPherson, A.L., Mouw, K.W., Rice, P.A. and Stark, M.S. (2009) Regulatory mutations in Sin recombinase support a structure-based model of the synaptosome. Molecular Microbiology, 74(2), pp. 282-298. (doi: 10.1111/j.1365-2958.2009.06756.x)

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Publisher's URL: http://dx.doi.org/10.1111/j.1365-2958.2009.06756.x

Abstract

The resolvase Sin regulates DNA strand exchange by assembling an elaborate interwound synaptosome containing catalytic and regulatory Sin tetramers, and an architectural DNA bending protein. The crystal structure of the regulatory tetramer was recently solved, providing new insights into the structural basis for regulation. Here we describe the selection and characterization of two classes of Sin mutations that respectively bypass or disrupt the functions of the regulatory tetramer. Activating mutations, which allow the catalytic tetramer to assemble and function independently at site I (the crossover site), were found at ~20% of residues in the N-terminal domain. The most strongly activating mutation (Q115R) stabilized a catalytically active synaptic tetramer <i>in vitro</i>. The positions of these mutations suggest that they act by destabilizing the conformation of the ground-state site I-bound dimers, or by stabilizing the altered conformation of the active catalytic tetramer. Mutations that block activation by the regulatory tetramer mapped to just two residues, F52 and R54, supporting a functional role for a previously reported crystallographic dimer-dimer interface. We suggest how F52/R54 contacts between regulatory and catalytic subunits might promote assembly of the active catalytic tetramer within the synaptosome.

Item Type:Articles
Status:Published
Refereed:Yes
Glasgow Author(s) Enlighten ID:Boocock, Dr Martin and Rowland, Dr Sally-Jane and Stark, Professor Marshall
Authors: Rowland, S.J., Boocock, M.R., MacPherson, A.L., Mouw, K.W., Rice, P.A., and Stark, M.S.
Subjects:Q Science > QH Natural history > QH345 Biochemistry
College/School:College of Medical Veterinary and Life Sciences > Institute of Molecular Cell and Systems Biology
Journal Name:Molecular Microbiology
ISSN:0950-382X
ISSN (Online):1365-2958
Published Online:08 June 2009

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