Interview to Steven B. Smith from Diego Salas Bruggink about stretched-DNA form

Interview to Steven B. Smith from Diego Salas Bruggink about stretched-DNA form

Fotografía tomada el 10 de febrero de 2018 por Christian A.M. Wilson en Steve garage

Steven B. Smith interest in single molecules started in 1988 when he was an electronic technician working at the University of Washington.  There he built equipment for the Genetics Department during the day and took graduate classes at night to earn a master’s degree in physics. For his thesis project, he studied how pulse-field gel electrophoresis can separate very large DNA molecules by size. He was surprised to find that he could see stained DNA molecules moving through agarose gel in an electric field by using a fluorescence microscope which belonged to the Genetics Department. Here is a paper that came from some of that work:

  1. B. Smith, P. D. Aldridge, and J. B. Callis, «Observation of Individual DNA Molecules Undergoing Gel Electrophoresis». Science 243:203-206 (1989) 

Entropic elasticity is the most interesting and obvious property of a polymer molecule as seen in a microscope. There the molecule stretches into a line under force but contracts to a ball when the force is relaxed.  Several theories existed about this force but there were no good experiments to test any theory. Then in 1990 he joined Carlos Bustamante’s laboratory at the University of New Mexico, and with graduate student Laura Finzi, they devised an experiment using magnetic beads to measure the entropic forces on single DNA molecules. They discovered that the popular entropic theory of that time (the Freely Jointed Chain model) could not predict entropic forces in DNA.

  1. B. Smith, L. Finzi, and C. Bustamante, «Direct Mechanical Measurements of the Elasticity of Single DNA Molecules by Using Magnetic Beads». Science 258,1122-1126 (1992)

Then John Marko and Eric Siggia and them developed a new equation for the Worm-Like Chain model to correctly fit their force data. 

  1. Bustamante J. Marko, E. Siggia, and S. B. Smith,  «Entropic Elasticity of l-Phage DNA», Science 265, 1599-1600 (1994)

What they did not mention in the 1992 paper was that they had seen some strange behavior of DNA which was not explained by either model. Under very high flow forces they saw the DNA molecules reversibly “over-stretch” far beyond their B-form contour lengths.  they wished to explore this stretch transition, but the magnetic beads they had available were too weak to measure the large forces required.

In 1993 Carlos Bustamante moved his laboratory to the University of Oregon where with graduate student Yujia Cui and him built an optical trap capable of exerting 100 pN force on a large (3-micron) refracting bead.  Inspiration for their instrument came from reading papers by Arthur Ashkin  (Phys Rev Lett v.24 #4 1970, and Optics Lett. v.11 #5 1986) and Tudor Buican (Proc. SPIE, 1990 and Cell Robotics Inc).  They then developed ways of measuring the force by changes in light momentum leaving the trap.  They first reported the overstretch transition as a poster abstract in Biophysical Journal vol. 68, A250 (1995).  Later a more complete study of the overstretch transition and measurements of single-stranded elasticity were sent to Science.

  1. B. Smith, Y. Cui and C. Bustamante, » Overstretching B-DNA: the Elastic Response of Individual Double-Stranded and Single-Stranded DNA Molecules», Science271, 795-798 (1996)

In that same issue, a group from France reported the same stretch transition at nearly the same force (70 pN versus 65 pN). Their measurements were made using fine glass needles which bent to indicate force.  They named the extended form of the molecule “S-DNA”.  Here is reference to their paper:

Philippe Cluzel, Anne Lebrun, Christoph Heller, Richard Lavery, Jean-Louis Viovy,

Didier Chatenay, François Caron, “DNA: An Extensible Molecule” Science 271, 792-794 (1996)