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Aspects of DNA Strand Exchange: Recombination Proteins and Model System Studies

Karolin Frykholm (Institutionen för kemi- och bioteknik, Fysikalisk kemi)
Göteborg : Chalmers University of Technology, 2010. ISBN: 978-91-7385-448-1.

DNA recombination is of fundamental importance to all living cells; it is part of the DNA repair machinery and a means to generate genetic diversity. DNA strand exchange, the exchange of strands between homologous DNA molecules, is the central reaction of the recombination process. The work presented in this Thesis has the aim of gaining insight into the mechanism of this reaction by investigating different aspects of DNA strand exchange. Structural studies of recombinase nucleoprotein filaments, which constitute the scaffold for the reaction in vivo, are reported together with investigations of artificial strand exchange in two different model systems.

The structures of active RecA and Rad51 nucleoprotein filaments have been studied by Site-Specific Linear Dichroism (SSLD), a spectroscopic approach based on linear dichroism in combination with molecular replacement of individual amino acids. In this Thesis it is shown how LD data of systematically engineered proteins can provide angular orientations for specific residues and how these coordinates can be used to build a structural model of the protein. From SSLD data of RecA it is concluded that the protein adopts similar structures in the initial and final states of strand exchange, indicating a static role for RecA during the reaction. The study of the human Rad51 protein illustrates how experimental data from SSLD can be successfully combined with theoretical molecular modelling. The outcome is a model structure of the protein in its active complex with DNA, the first detailed structure reported so far for the complete human Rad51 protein.

This Thesis reports on artificial strand exchange catalysis aided by cationic lipid vesicles and it is shown that DNA opens up in a zipper-like fashion, which facilitates strand exchange. It is further concluded that the exchange mechanism on the liposome surface is fundamentally different from that in bulk solution. Non-ionic catalysis has been investigated by the use of PEG to induce molecular crowding and provide possibilities for hydrophobic interactions. PEG accelerates strand exchange dramatically and the results emphasize the importance of hydrophobic interactions between DNA and its environment for the dynamic behaviour of DNA strands.

Nyckelord: DNA strand exchange, RecA, Rad51, Site-Specific Linear Dichroism, cationic liposome, molecular crowding

Denna post skapades 2010-10-01. Senast ändrad 2013-09-25.
CPL Pubid: 127152


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Institutioner (Chalmers)

Institutionen för kemi- och bioteknik, Fysikalisk kemi (2005-2014)


Biofysikalisk kemi

Chalmers infrastruktur

Relaterade publikationer

Inkluderade delarbeten:

Conserved conformation of RecA protein after executing the DNA strand-exchange reaction. A site-specific linear dichroism structure study

Enhanced DNA strand exchange on positively charged liposomes

Structure of human Rad51 protein filament from molecular modeling and site-specific linear dichroism spectroscopy

Mechanism of DNA Strand Exchange at Liposome Surfaces Investigated Using Mismatched DNA


Datum: 2010-10-29
Tid: 13:15
Lokal: KC-salen, Kemigården 4, Chalmers, Göteborg
Opponent: Professor Andrzej Stasiak, University of Lausanne, Schweiz

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Doktorsavhandlingar vid Chalmers tekniska högskola. Ny serie 3129