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42555-G9
Translocation of Structured Polymers through Nanopores
The goal of this project is to qualitatively and quantitatively model the translocation of structured molecules, specifically RNA and single-stranded DNA, through nanopores. These nanopores are so small that only single-stranded RNA and DNA can pass through them - any Watson-Crick base pairs between bases of the molecules have to be broken in order for the molecule to pass through the pore. Thus, the dynamics of translocation of structured biomolecules through nanopores provides information about base-pairing dynamics of these molecules - a topic that in and of itself is of great interest since many biological processes are greatly effected by base pairing dynamics.
One interesting phenomenon that I have been studying in this second year of support by the Petroleum Research Fund is the phenomenon of anomalous scaling of translocation times. Naively, one would expect that for two molecules, one of which is twice as long as the other, the translocation time of the longer one is also twice the translocation time of the shorter one. Thus, deviations from this naive expectation are interesting. For unstructured molecules such deviations are known to arise from entropic effects. These entropic effects do not play any role for translocation of structured molecules and thus the question arises if anomalous dynamics can be present for structured molecules as well. To study this, I am collaborating with specialists on anomalous dynamics, Yariv Kafri (Technion, Haifa, Israel) and Julius Lucks (Cornell), as well as with my long term collaborator Ulrich Gerland (Universitaet zu Koeln, Germany). While this work is still in progress, we have identified two possible driving forces for anomalous dynamics: the fluctuations in the energy landscape due to the sequence variations along the molecule and the average shape of this energy landscape in and of itself. We have strong numerical evidence for anomalous dynamics caused by the average shape of the energy landscape. The data on the influence of fluctuations is less pronounced and we are currently studying this aspect in more detail before publishing this work.
Another significant effort was to implement a quantitatively correct energy model for the base pairing dynamics and applying it to published data on the translocation of DNA hairpins (the simplest base pairing structure possible) from the Meller laboratory. While a significant amount of effort has been devoted to this sub-project, it has not yielded the results hoped for, namely a microscopic understanding of translocation dynamics. One serious problem was the staffing of the project. A graduate student worked for two quarters on this project but it turned out that while he was technically quite proficient he was lacking the drive to actually study the scientific problems at hand and do any more than what he was directly instructed to do. Thus, a new student had to be trained to continue the project. The other problem emerging in this line of the research is a discrepancy of several orders of magnitude between our simulation results and the experiments. It appears as if the presence of the pore changes the thermodynamics of DNA dissociation in some dramatic way - however, in spite of very intensive and frequent discussions with experimentalist Amit Meller all effects of the pore that we have considered so far lead to a change of the translocation time in the opposite direction of what is observed. We continue to investigate this issue vigorously.
A third sub-project does not involve a nanopore per se but still attempts to understand the reaction of nucleic acids to external constraints. More precisely, we are studying the ability of short double stranded DNAs to cyclize. While this cyclization ability for longer molecules is well understood on the basis of pure polymer physics models, it is for short molecules expected to be driven by the formation of local unpaired regions - just like the unpairing when translocating through a nanopore - and thus to depend in a very specific way on the sequence of the molecule and on external parameters such as temperature. Graduate student Bob Forties and I developed a quantitative model of this phenomenon. Currently, Bob Forties is verifying this model experimentally in the laboratory of my colleague Michael Poirier. For the sequences studied so far, we find excellent agreement between theory and experiment.
The results of this research have been presented in several invited talks, e.g., at a workshop on the Physical and Chemical foundations of Bioinformatics Methods in Dresden, Germany, a seminar at Rochester University Medical School, and summer schools in Singapore as well as St. Etienne du Tinees, France. Especially the latter allowed many students to benefit from this work in addition to the actual students in my laboratory to whom this project provided insight into truly interdisciplinary work.
Acknowledgment is made to the Donors of the American Chemical Society Petroleum Research Fund for support of this research.
