Trial Lecture – time and place
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Adjudication committee
- First opponent: Professor Marit Sletmoen, Department of Biotechnology and Food Science, Norwegian University of Science and Technology
- Second opponent: Associate Professor Sebastian Deindl, Institute of Cell and Molecular Biology, Uppsala University, Sweden
- Third member and chair of the evaluation committee: Associate Professor Tor Erik Rusten, Institute of Clinical Medicine, University of Oslo
Chair of the Defence
Professor Philippe Collas, Institute of Basic Medical Sciences, University of Oslo
Principal Supervisor
Researcher Bjørn Dalhus, Institute of Clinical Medicine, University of Oslo
Summary
To combat the destructive effects of damage to DNA, different repair mechanisms play central roles. DNA is repaired by the activity of several classes of DNA repair proteins which scan the DNA to locate the damage, then repairing the lesions. Despite decades of intensive research and great progresses in identifying the structures and biochemical functions of DNA repair proteins, a detailed understanding of the molecular mechanisms by which they search for and recognize errors in DNA remains elusive.
The overall aim of this study was to characterize and compare the DNA scanning mechanisms of selected DNA repair proteins at the single-molecule level, and to look for a possible relationship between scanning behavior and structural features of these proteins. In our single-molecule assay, a 4 µm long fragment of DNA is elongated by attaching it to a microscope coverslip at one end and to a polystyrene bead held in an optical trap at the other end. The elongated DNA is exposed to fluorescently labelled proteins and their interaction with DNA is recorded as trajectories of movement of proteins along the DNA, with images taken at rates up to 130 Hz.
From the analysis of these trajectories we find that during DNA scanning, the enzyme EndoV switches between three different scanning modes which can be generally classified as helical sliding, hopping and base interrogation. We also show that the highly conserved wedge motif in the structure of this protein plays a central role in switching DNA scanning into base interrogation mode. In contrast to DNA glycosylases such as human OGG1, the Heat-Like Repeat (HLR) protein AlkD is known as a DNA glycosylase whose activity does not depend on DNA bending or base-flipping. We show that DNA scanning by AlkD resembles a single-mode random walk contrary to the multi-mode scanning utilized by the other DNA repair proteins in this study. This result resonates well with the lack of base flipping in AlkD.
Additional information
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