Methods of single-molecule energy landscape reconstruction with optical traps

Optical traps facilitate measurement of force and position as single molecules of DNA, RNA, or protein are unfolded and refolded. The effective energy landscape of a biomolecule can be reconstructed from the force and position data, providing insight into its structure and regulatory functions. We have developed new experimental and analytical methods to reconstruct energy landscapes by taking advantage of the harmonic constraint of an optical trap. We demonstrate the effectiveness of these methods using a model DNA hairpin and then apply these methods to study problems of practical biophysical interest. CCR5 mRNA has been demonstrated to stimulate -1 programmed ribosomal frameshifting and we measure its structural properties. We measure the binding energy of a GA/AG tandem mismatch, one of many mismatches with unusual properties. We use our single-molecule methods to reproduce bulk measurements of the nearest-neighbor DNA base-pair free energy parameters and we consider possible refinements to the model. We also study an alternative method of measuring energy landscapes, Dynamic Force Spectroscopy (DFS), and conduct experiments on DNA quadruplexes to demonstrate the effectiveness of DFS with optical traps. Finally, we develop theory to elucidate the role of noise in optical trap measurements of energy landscapes.

Department dc:contributor.department

Physics

Year dc:date.issued

2012

Author dc:creator

• de Messieres, Michel Escande

Advisor dc:contributor.advisor

• La Porta, Arthur

Handle dc:identifier.uri

http://hdl.handle.net/1903/12737

OAI identifier oai:identifier

oai:drum.lib.umd.edu:1903/12737