Difference Topology: extending the Tangle Model
Jointly with John Luecke and Isabel Darcy, I have analyzed data from novel difference topology experiments to unveil the structure of the Mu transpososome. The technique was developed in Rasika Harshey’s and Makkuni Jayaram’s lab, and the experiments were done by Shailja Pathania.
The purpose in difference topology experiments is to shed light on the topological structure of a stable protein/DNA complex (such as the Mu transpososome) by using a known site-specific recombinase to create DNA knots or links when given the complex as a substrate. The resulting knots/links contain the information needed to understand the whole complex. Analyzing difference topology experiments requires extending the tangle model, especially since the complex to be analyzed may be better modeled by n-string tangles, where n is greater than 2.
The Mu transpososome
Transposable elements, also called mobile elements, are fragments of DNA able to move along a genome by a process called transposition. Mobile elements play an important role in the shaping of a genome, and they can impact the health of an organism by introducing genetic mutations. Of special interest is that transposition is mechanistically very similar to the way certain retroviruses, including HIV, integrate into their host genome.
Bacteriophage Mu is a system widely used in transposition studies due to the high efficiency of Mu transposase. The MuA protein performs the first steps required to transpose the Mu genome from its starting location to a new DNA location. MuA binds to specific DNA sequences which we refer to as attL and attR sites (named after Left and Right attaching regions). A third DNA sequence called the enhancer (E) is also required to assemble the Mu transpososome. The Mu transpososome is a very stable complex consisting of 3 segments of double- stranded DNA captured in a protein complex. In our paper we are interested in studying the topological structure of the DNA within the Mu transpososome.
Look at the preprint: Darcy et al, submitted