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Using lasers to manipulate and visualize single DNA molecules: DNA repair in action

Event Details

Event Dates: 

Monday, June 27, 2016 - 4:00pm

Seminar Location: 

  • JILA X317

Speaker Name(s): 

Erwin Peterman

Speaker Affiliation(s): 

University of Amsterdam
Seminar Type/Subject

Scientific Seminar Type: 

  • JILA Public Seminar

Event Details & Abstract: 

Reception at 3:30 in HBar

In my lab, we use lasers to simultaneously manipulate and visualize biomolecules, at the single-molecule level. Over the years we have developed and applied a unique combination of multi-beam optical tweezers and (single-molecule) fluorescence microscopy. The optical tweezers allows holding, extending, applying tension to and measuring force on DNA molecules, while fluorescence microscopy can used, at the same time, to probe the structure of the DNA or proteins binding to it. We have successfully applied this combination of tools to study DNA structure and mechanical stability, DNA compaction, replication, transcription and repair. In my presentation, I will explain the background of our combined approach and I will focus on a recent application on the role of the proteins XRCC4 and XLF in Non-Homologous End Joining (NHEJ), an essential DNA-repair mechanism that mends DNA double-strand breaks. We use our approach combining optical tweezers and fluorescence microscopy to characterize the interaction of XRCC4-XLF complexes with individual DNA molecules. We find that XRCC4-XLF complexes stably bind DNA but diffuse rapidly, undergoing continuous rearrangements and switching between static and dynamic binding modes. By simultaneous manipulation of two individual DNA molecules we find that XRCC4-XLF complexes form intermolecular DNA bridges. XRCC4-XLF bridges are mobile and resist to remarkably high pulling forces. These results shed new light on the dynamics of NHEJ.

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