Nano-biology: Single-molecule DNA Nanomanipulation and Single-molecule Fluorescence

Program: Nanobiology
Department: Chemistry and Chemical Biology
Collaborator: Terence Strick, Department of Physics, University of Paris
By monitoring the overall extension of a mechanically stretched, negatively supercoiled, single DNA molecule containing a single target site, we are able to observe the gain of ~1 positive supercoil, or loss of ~1 negative supercoil, associated with protein-induced unwinding of ~10 base pairs of the target site.1 The assay enables us to define--at the single-molecule level, in real time the extent of unwinding, the kinetics of unwinding, and the lifetime and dynamics of the unwound state. In addition, the assay enables us to define effects of DNA sequence, supercoiling, temperature, and small molecule modulators.

We are working to develop single-molecule-nanomanipulation and combined single-molecule-nanomanipulation/single-molecule-fluorescence approaches for systematic structural and mechanistic analysis of nucleic-acid processing enzymes, to automate the approaches, and to apply the approaches to drug discovery.

  1. Proc. Natl. Acad. Sci. USA, 101:4776-4780 (2004).