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Capture of a Single Molecule in a Nanocavity Li-Qun Gu, Stephen Cheley, Hagan Bayley

Capture of a Single Molecule in a Nanocavity Li-Qun Gu, Stephen Cheley, Hagan Bayley Department of Medical Biochemistry and Genetics, Texas A&M University Science 2001 (Volume 291, page 636). Outline. Why nanothingies are interesting. Other Approaches Schematic Outline Implementation

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Capture of a Single Molecule in a Nanocavity Li-Qun Gu, Stephen Cheley, Hagan Bayley

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  1. Capture of a Single Molecule in a Nanocavity Li-Qun Gu, Stephen Cheley, Hagan Bayley Department of Medical Biochemistry and Genetics, Texas A&M University Science 2001 (Volume 291, page 636)

  2. Outline • Why nanothingies are interesting. • Other Approaches • Schematic Outline • Implementation • Results • Future Possibilities

  3. Why Nanothingies are Interesting • Diffusion length is 1 m2/ms or 1nm2/ns giving a mean free path of 0.1 Angstroms or so.(Andersen, O.S., S.W. Feldberg “Journal of Physical Chemistry” Vol 100 p4622-4629) • Study the process of biological filtration (kidney nephrons, liver, thyroid, lymphic system, nasal and other mucosal membranes etc.) • Improve electrophoresis, molecular exclusion chromatography and other filtration techniques. • Develop new chemical techniques utilizing unique catalysis of molecules bound to nanopores • Develop sensors for binding to nanopores (eg. Surface Treated BiaCORE Surface Plasmon Resonance Detector) • Study molecules as they pass through nanopores - DNA Sequencing • Time Resolved studies of binding, folding and other molecular events.

  4. Sequencing DNA through a NanoPore The passage of a polymer through an small pore should alter the conductivity of the pore. Ideally, each monomer will have different binding characteristics in the pore. The current through the pore amplifies the single molecule event. “Microsecond Time-Scale Discrimination Among Polycytidylic Acid, Polyadenylic Acid, and Polyuridylic Acid as Homopolymers or as Segments witting Single RNA molecules” Mark Akeson, Daniel Branton, John J. Kasianowicz, Eric Brandin, David W. Deamer. Biophysical Journal 1999 Volume 77 p3227-3233

  5. Rapid nanopore discrimination between single polynucleotide moleculesAmit Meller, Lucas Nivon, Eric Brandin, Jene Golvchenko, Daniel BrantonDepartment of Physics, Harvard UniversityPNAS 2001 : vol 97. p1079-1084 • Fig. 5. The individual events are identified, on the basis of tD alone, as traversal of a molecule of poly(dA)100 or a molecule of poly(dC)100. - Study passage of poly-adenosine and poly-cytosine residues (100 bp) through an -Hemolysin channel - Find can clearly distinguish sequences.

  6. Nanotubule-Based Molecular Filtration Membranes Kshama B. Jirage, John C. Hulteen, Charles R. Martin Science 1997 Vol 278 p655. Figure 1. Schematic illustrations of the shapes of the Au nanotubes that we obtained by doing the electrodeless Au plating (9) at (A) pH = 10 and (B) pH = 12. The higher pH causes bottleneck tubules. The tubules plated at the lower pH also have some of this bottleneck character [see (11)]. Hence, the depictions in both (A) and (B) are approximate and serve to illustrate the conceptual differences between the two types of nanotubules investigated here.

  7. Schematic Outline • Want a cavity in which a guest molecule, G, can be trapped. • Put two plugs in an existing pore. • G binds but can pass through the top plug. • G cannot get through bottom plug. • Monitor G’s position by current through pore.

  8. Cyclo-Dextran Structure CD : R=OH ; s7CD : R = 0S03 Alpha HemoLysin Both figures taken from L-Q. Gu et al. PNAS 97:3959 (2000) • Alpha Hemolysin is a seven-subunit protein (293) pore from staphylococcal aureus. Water soluble and added from cis side. Forms a beta barrel of 15 Angstroms diameter or so. • CDs are 7-fold cyclic sugars. • Mutate Alpha Hemolysin to bind CDs when added from trans side

  9. Proof of Binding of Two Plugs To demonstrate that plugs (CD and S7CD) both work as expected, molecules G1 (1,3 adamantane dicarboxylic acid) and G2 (1-Adamantane carboxamide) were added to the cis and trans compartments respectively (Vtrans = -80mv)

  10. Demonstration that Plugs work as Expected (test done at Vtrans = -80mV)

  11. The Whole-Shebang in Action Test done at Vtrans = 80mV

  12. Future Advances in NanoCavities • Covalently bind plug/converters in place. • Develop plug/converters with useful chemical selectivity instead of picking Guest molecule, G, to fit the plug. • Alter the wall coating of the beta-barrel. • Place the nanocavity on a supported or polymerized membrane • Add binding sites to the nanopore • Include fluorescent labels in the nanopore.

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