Dr. Eyal Nir May, 2015
CURRICULUM VITAE
• Personal Details Name: Eyal Nir Date and place of birth: 06-08-1971 Address and telephone number at work: Ben-Gurion University of the Negev, Beer Sheva, Israel; Office: 972(0)8-6428474, Lab: 972(0)8-6428472 Home Address: HaAtad Street, #11, Apartment #14, Tel-Aviv, 66843, Israel, Phone: 972(0)50-6994468
• Education B.Sc. (1994-1997) Hebrew University, Jerusalem, Israel, Chemistry
M.Sc. (1998-1999) Hebrew University, Jerusalem, Israel, Physical-Chemistry Name of advisor: Prof. Mattanjah de Vries Title of the Thesis: Building Blocks of DNA, Gas-Phase Research
Ph.D. (1999-2003) Hebrew University, Jerusalem, Israel, Physical-Chemistry Name of advisor: Prof. Mattanjah de Vries Title of the Thesis: Building Blocks of DNA, Gas-Phase Research
• Employment History 10/2008 – Current: Senior Lecturer, Department of Chemistry, Ben-Gurion University of the Negev 2003 – 2008: Research Associate (Post-doctorate), Department of Chemistry and Biochemistry, University California Los-Angeles (UCLA), USA 2001 – 2002: Researcher, Department of Physical-Chemistry, University of California Santa Barbara (UCSB), USA 1997 – 2001: Teaching Assistant, Chemistry Department, Hebrew University, Israel
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• Professional Activities Ad-hoc reviewer for peer-reviewed journals Science, Journal of the American Chemical Society, Nano Letters, Angewandte Chemie, ACS-Nano, The Journal of Physical Chemistry (Letters), Physical Chemistry Chemical Physics, Molecules, Nanoscale, PLOS ONE, Methods, Scientific Reports, A
• Educational Activities (a) Courses Taught 1. Physical-Chemistry-I course for 2nd year undergraduate pharmacist (2009, 2011, 2012, 2013, 2014, 2015), biologist, geologists, computer science and health science students (2010, 2012, 2013, 2014, 2015) 2. Physical-Chemistry-II course for 3nd year undergraduate pharmacist (2010, 2011, 2012, 2013, 2014, 2015) 3. Physical-Chemistry-Advanced course for 3nd year undergraduate chemists together with Prof. Pines (2009, 2010, 2013, 2014) 4. Physical-Chemistry Lab - A for 2nd year undergraduate chemists (2009, 2011) 5. Physical-Chemistry Lab - B for 3nd year undergraduate chemical engineers (2011, 2014, 2015) 6. Physical-Chemistry Advance Lab - A for 3nd year undergraduate chemists (2010) 7. Condensed-Phase Course for 3nd year undergraduate chemists (2011, 2014 – with Dr. Visoly, 2015) 8. Physical-Chemistry Seminar Coordinator for graduate students (2011) 9. Departmental Seminar Coordinator for graduate students (2013) 10. General and Analytical Chemistry Lab for 1st year undergraduate chemists (2013)
(b) Research Students Graduated: Rula Masoud (M.Sc. 2011) Toma Tomov (M.Sc. 2011, continued to doctorate studies in our group) Yaron Berger (M.Sc. 2014, continued to doctorate studies in our group) Roman Tsukanov (Ph.D. 2014, continued post-doctorate research in our group) Noa Plavner (Ph.D. 2015)
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Current Students and Post-doctorate Toma Tomov, Doctorate studies, since 2011 – NEGEV Fellowship, 2011 Miran Liber, Doctorate studies, since 2011 – MILGAT DAROM Fellowship, 2011 Yaron Berger, Doctorate studies, since 2014 Roman Tsukanov, Post-doctorate, since 2014 Menahem Pirchi, Post-doctorate, since 2014 Michal Pellach, Post-doctorate, since 2015 Dinesh Khara, Post-doctorate, since 2015
Other Alumni Tapasi Sen, Post-doctorate, 2013
• Awards, Citations, Honors, Fellowships (a) Honors, Citation Awards Intel-Dean Prize for Excellent Ph.D. research, 2000, Hebrew University (4000$)
(b) Fellowships 1. DFG (Deutsche Forschungs Gemeinschaft), 1999-2002, Ph.D. Fellowshiop (~45,000$) 2. HFSP (Human Frontier Science Program), 2003-2006, Post-doctorate Fellowship (~150,000$) 3. ALON Fellowship, 2009, Young Faculty in Israel (~230,000$)
• Scientific Publications - Information is from ISI WEB OF SCIENCE Peer-reviewed papers: 29 Book chapters: 2 Total number of scientific citations, without self-citations: 1857 H-Index: 20 (out of 21 peer-reviewed papers from Ph.D. and post-doctorate studies)
(a) Peer-Reviewed Journals (*From BGU)
1. Nir, E.S; Grace, L.T; Brauer, B.C; de Vries, M. S.PI, "REMPI spectroscopy of jot-cooled guanine", Journal of the American Chemical Society, 1999, 121, 4896-4897. (IF = 11.444; JR = 10/148; Q1; Citations = 103) 2. Nir, E.S; Hunziker, H. E.PI; de Vries, M. S.PI, "Fragment-free mass spectrometric analysis with jet cooling VUV photoionisation", Analytical Chemistry, 1999, 71, 1674- 1678. (IF = 5.825; JR = 4/76; Q1; Citations = 41) Dr. Eyal Nir page 4
3. Cohen, R.S; Brauer, B.C; Nir, E.S; Grace, L.T; de Vries, M. S.PI, "Resonance-enhanced multiphoton ionization spectroscopy of dipeptides", Journal of Physical Chemistry A, 2000, 104, 6351-6355. (IF = 2.775; JR = 50/136; Q2; Citations = 83) 4. Nir, E.S; Imhof, P.S; Kleinermanns, K.PI; de Vries, M. S.PI, "REMPI spectroscopy of laser desorbed guanosines", Journal of the American Chemical Society, 2000, 122, 8091-8092. (IF = 11.444; JR = 10/148; Q1; Citations = 54) 5. Nir, E.S; Kleinermanns, K.PI; de Vries, M. S.PI, "Pairing of isolated nucleic-acid bases in the absence of the DNA backbone", NATURE, 2000, 408, 949-951. (IF = 42.351; JR = 1/55; Q1; Citations = 183) 6. Nir, E.S; Janzen, C.S; Imhof, P.S; Kleinermanns, K.PI; de Vries, M. S.PI, "Guanine tautomerism revealed by UV-UV and IR-UV hole burning spectroscopy", Journal of Chemical Physics, 2001, 115, 4604-4611. (IF = 3.122; JR = 8/33; Q1; Citations = 145) 7. Nir, E.S; Kleinermanns, K.PI; Grace, L.T; de Vries, M. S.PI, "On the photochemistry of purine nucleobases", Journal of Physical Chemistry A, 2001, 105, 5106-5110. (IF = 2.775; JR = 50/136; Q2; Citations = 134) 8. Plutzer, C.S; Nir, E.S; de Vries, M. S.PI; Kleinermanns, K.PI, "IR-UV double-resonance spectroscopy of the nucleobase adenine", Physical Chemistry Chemical Physics, 2001, 3, 5466-5469. (IF = 4.198; JR = 33/136; Q1; Citations = 90) 9. Nir, E.S; de Vries, M. S.PI, "Fragmentation of laser-desorbed 9-substituted adenines", International Journal of Mass Spectrometry, 2002, 219, 133-138. (IF = 2.227; JR = 13/33; Q2; Citations = 5) 10. Nir, E.S; Janzen, C.S; Imhof, P.S; Kleinermanns, K.PI; de Vries, M. S.PI, "Pairing of the nucleobases guanine and cytosine in the gas phase studied by IR-UV double- resonance spectroscopy and ab initio calculations", Physical Chemistry Chemical Physics, 2002, 4, 732-739. (IF = 4.198; JR = 33/136; Q1; Citations = 95) 11. Nir, E.S; Janzen, C.S; Imhof, P.S; Kleinermanns, K.PI; de Vries, M. S.PI, "Pairing of the nucleobase guanine studied by IR-UV double-resonance spectroscopy and ab initio calculations", Physical Chemistry Chemical Physics, 2002, 4, 740-750. (IF = 4.198; JR = 33/136; Q1; Citations = 79) 12. Nir, E.S; Muller, M.S; Grace, L.T; de Vries, M. S.PI, "REMPI spectroscopy of cytosine", Chemical Physics Letters, 2002, 355, 59-64.(IF = 1.991; JR= 77/136; Q3; Citations = 98) 13. Nir, E.S; Plutzer, C.S; Kleinermanns, K.PI; de Vries, M. S.PI, "Properties of isolated DNA bases, base pairs and nucleosides examined by laser spectroscopy", European Physical Journal D, 2002, 20, 317-329. (IF = 1.398; JR = 40/82; Q2; Citations = 148) 14. Nir, E.S; Hunig, I.S; Kleinermanns, K.PI; de Vries, M. S.PI, "The nucleobase cytosine and the cytosine dimer investigated by double resonance laser spectroscopy and ab initio calculations", Physical Chemistry Chemical Physics, 2003, 5, 4780-4785. (IF = 4.198; JR = 33/136; Q1; Citations = 55) 15. Plutzer, C.S; Hunig, I.S; Kleinermanns, K.PI; Nir, E.S; de Vries, M. S.PI, "Pairing of isolated nucleobases: Double resonance laser spectroscopy of adenine-thymine", Chemphyschem, 2003, 4, 838-842. (IF = 3.360; JR = 7/33; Q1; Citations = 62) Dr. Eyal Nir page 5
16. Nir, E.S; Hunig, I.S; Kleinermanns, K.PI; de Vries, M. S.PI, "Conformers of guanosines and their vibrations in the electronic ground and excited states, as revealed by double-resonance spectroscopy and ab initio calculations", Chemphyschem, 2004, 5, 131-137. (IF = 3.360; JR = 7/33; Q1; Citations = 32) 17. Abo-Riziq, A.S; Grace, L.T; Nir, E.S; Kabelac, M.S; Hobza, P.PI; de Vries, M. S.PI, "Photochemical selectivity in guanine-cytosine base-pair structures", Proceedings of the National Academy of Sciences of the United States of America, 2005, 102, 20-23. (IF = 9.809; JR = 4/55; Q1; Citations = 134) 18. Jager, M.PD; Nir, E.PD; Weiss, S.PI, "Site-specific labeling of proteins for single- molecule FRET by combining chemical and enzymatic modification", Protein Science, 2006, 15, 640-646. (IF = 2.861; JR = 146/291; Q3; Citations = 27) 19. Nir, E.PD; Michalet, X.C; Hamadani, K. M.S; Laurence, T. A.S; Neuhauser, D.C; Kovchegov, Y.C; Weiss, S.PI, "Shot-noise limited single-molecule FRET histograms: Comparison between theory and experiments", Journal of Physical Chemistry B, 2006, 110, 22103-22124. (IF = 3.377; JR = 39/136; Q2; Citations = 119) 20. Kong, X. X.S; Nir, E.PD; Hamadani, K.S; Weiss, S.PI, "Photobleaching pathways in single-molecule FRET experiments", Journal of the American Chemical Society, 2007, 129, 4643-4654. (IF = 11.444; JR = 10/148; Q1; Citations = 35) 21. Majumdar, D. S.S; Smirnova, I.T; Kasho, V.T; Nir, E.PD; Kong, X.S; Weiss, S.PI; Kaback, H. R.PI, "Single-molecule FRET reveals sugar-induced conformational dynamics in LacY", Proceedings of the National Academy of Sciences of the United States of America, 2007, 104 (31), 12640-12645. (IF = 9.809; JR = 4/55; Q1; Citations = 91) *22. Kovchegov, Y.PI; Meredith, N.S; Nir, E.PI, "Occupation times and Bessel densities", Statistics & Probability Letters, 2010, 80, 104-110. (IF = 0.531; JR = 96/119; Q4; Citations = 0) *23.Tomov, T. E.S; Tsukanov, R.S; Masoud, R.S; Liber, M.S; Plavner, N.S; Nir, E.PI; "Disentangling subpopulations in single-molecule FRET and ALEX experiments with photon distribution analysis", Biophysical Journal, 2012, 102 (5), 1163-1173. (IF = 3.832; JR = 18/74; Q1; Citations = 11) *24. Masoud, R.S; Tsukanov, R.S; Tomov, T. E.S; Plavner, N.S; Liber, M.S; Nir, E.PI, "Studying the Structural Dynamics of Bipedal DNA Motors with Single-Molecule Fluorescence Spectroscopy", ACS Nano, 2012, 6(7), 6272-6283. (IF = 12.033; JR = 9/148; Q1; Citations = 8) *25. Tomov, T. E.S; Tsukanov, R.S; Liber, M.S; Masoud, R.S; Plavner, N.S; Nir, E.PI, "Rational Design of DNA Motors: Fuel Optimization through Single-Molecule Fluorescence", Journal of the American Chemical Society, 2013, 135, 11935. (IF = 11.444; JR = 10/148; Q1; Citations = 4) *26. Tsukanov, R.S; Tomov, T. E.S; Masoud, R.S; Drory, H.S; Plavner, N.S; Liber, M.S; Nir, E.PI, "Detailed Study of DNA Hairpin Dynamics Using Single-Molecule Fluorescence Assisted by DNA Origami", Journal of Physical Chemistry B, 2013, 117, 11932. (IF = 3.377; JR = 39/136; Q2; Citations = 3) *27. Tsukanov, R.S; Tomov, T. E.S; Berger, Y.S; Liber, M.S; Nir, E.PI, "Conformational Dynamics of DNA Hairpins at Millisecond Resolution Obtained from Analysis of Single-Molecule FRET Histograms", Journal of Physical Chemistry B, 2013, 117, 16105. (IF = 3.377; JR = 39/136; Q2; Citations = 2) Dr. Eyal Nir page 6
*28. Tsukanov, R.S; Tomov, T. E.S; Liber, M.S; Berger, Y.S; Nir, E.PI, "Developing DNA Nanotechnology Using Single-Molecule Fluorescence", Accounts of Chemical Research, 2014, 47 (6), 1789-1798. (IF = 24.348; JR = 3/148; Q1; Citations = 0) *29. Liber, M.S; Tomov, T. E.S; Tsukanov, R.S; Berger, Y.S; Nir, E.PI, "A Bipedal DNA Motor that Travels Back and Forth between Two DNA Origami Tiles", Small, 10.1002/smll.201402028, 08/2014. (IF = 7.514; JR = 15/148; Q1; Citations = 0)
(b) Book Chapters 30. Nir E, Jager M, Weiss S, “Alternating Laser Excitation spectroscopy of freely diffusing single molecule: application to bio-molecular structure, dynamics and interaction”, Chapter 2 in “Nano Biophotonics” volume 3, ELSEVIER. (Masuhara H, Kawata S and Tokunaga F, Eds. 2007). 31. Kapanidis AN, Heilemann, Margeat E, Kong X, Nir E, Weiss S, “Alternating-laser excitation of single molecules”, Chapter 2 in "Laboratory manual for single-molecule studies", CSHL Press, Cold Spring Harbor, NY (Taekjip Ha and Paul Selvin, Eds. 2008).
• Lectures and Presentations at Meetings and Invited Seminars (a) Invited and Plenary Lectures at Conferences/Meetings 1. “Listening to the Sounds of DNA, Gas-Phase Spectroscopy of DNA Building Blocks”, Ion Chemistry and Mass Spectrometry, Lake Arrowhead, CA, 01/2002, (Invited Talk) 2. “Fast Conformational Dynamics of Freely Diffusing DNA-Hairpin Extracted From spFRET Histogram Shape Analysis”, 50th Biophysical Society, Salt-Lake, Utah, US, 02/2006, (Talk) 3. “Fast Structural Dynamics of Freely Diffusing DNA-Hairpin Extracted From sp-FRET Histogram Shape Analysis”, Optical Spectroscopy of Bimolecular Dynamics II, Eilat, Israel, 03/2006, (Talk) 4. “Protein and DNA Structure and Dynamics By Single-Molecule Spectroscopy ALEX- FRET”, Single Molecule workshop, PicoQuant, Berlin, Germany, 09/2007, (Key Note Speaker) 5. “Determining Conformational Dynamics of Proteins and DNA by Single Molecule ALEX-FRET Experiments”, 52th Biophysical Society, Long-Beach, CA, USA, 02/2008, (Talk) 6. “Doing DNA-Nanotechnology Using Single Molecule Fluorescence Spectroscopy“, The 76th Israeli Chemical Society, Tel-Aviv, Israel, 01/2011, (Invited Talk) 7. “Distinguishing between Subpopulations in Single-Molecule FRET and ALEX Experiments using Photon Distribution Analysis”, The 17th Single-Molecule workshop, PicoQuant, Berlin, Germany, 09/2011, (Talk) Dr. Eyal Nir page 7
8. “Towards a Reliable and Useful Non-Autonomous DNA-Motors“, The Israeli Biophysical Society, Tel-Aviv, Israel, 10/2011, (Invited Talk) 9. “Towards a Reliable and Useful DNA-Motors and Devices: The Single-Molecule Fluorescence Strategy”, The First HUJI-UK Nano-Life Workshop, Jerusalem, Israel 06/2012, (Invited Talk) 10. “Towards Reliable DNA-Motors and Devices”, The German Biophysical Society, Gottingen, Germany, 09/2012, (Talk) 11. The 10th annual FNANO conference, Utah, US, 04/2013, (Invited Talk), (This is the most important conference in the field of DNA-Nanotechnology, I could not attend, my student Tommy Tomov presented instead) 12. “Toward Fast and Efficient DNA-Based Molecular Machines”, Single-Molecule Biophysics, Neve-Ilan, Israel, 05/2013, (Invited Talk) 13. “Rational Design and Operation of Fast and Efficient DNA-Based Nano-Machines using Single-Molecule Fluorescence and Microfluidics Technology”, Chinanano, China, Beijing, 08/2013, (Talk) 14. “Fast and Efficient DNA-Based Molecular Machines”, Israeli Vacuum Society, Herzeliya, Israel, 09/2013, (Invited Talk) 15. “Fast and Efficient DNA-Based Molecular Machines Assisted by Microfluidics and Single-Molecule Fluorescence”, Sonderforschungsbereic-1023, Munich, Germany, 12/2013, (Invited Talk) 16. “Fast and Efficient DNA-Based Molecular Machines Assisted by Microfluidics and Single-Molecule Fluorescence”, Israeli Chemical Society, Tel-Aviv, Israel, 02/2014, (Invited Talk) 17. “Fast and Efficient DNA Based Molecular Motors Assisted by Microfluidics and Single-Molecule Fluorescence”. International Meeting of the Physics of Living Systems, Munich, Germany, 07/2014, (Invited Talk) 18. “Combining Microfluidics Technology and Single-Molecule Fluorescence to Operate and Study DNA Molecular Machines”, The 20th Single-Molecule workshop, PicoQuant, Berlin, Germany, 09/2014, (Talk) 19. “Computer Controlled DNA-Based Molecular Machines Assisted by Microfluidics and Single-Molecule Fluorescence”, Special Secession on Single-Molecule Fluorescence for the Honor of Nobel Laureate William. E. Moerner, Israeli Chemical Society, Tel- Aviv, Israel, 02/2015, (Invited Talk) 20. “Computer Controlled DNA-Based Molecular Machines Assisted by Microfluidics and Single-Molecule Fluorescence”, Israel Society for Microscopy, Bar Ilan, Israel, 05/2015, (Invited Talk) Dr. Eyal Nir page 8
21. “Computer Controlled DNA-Based Molecular Machines Assisted by Microfluidics and Single-Molecule Fluorescence”, Super Resolution Workshop. Sde-Boker, Israel, 05/2015, (Talk)
(b) Presentation of Papers at Conferences/Meetings (Posters) 22. “DNA Building Blocks; Gas-Phase Research”, Isolated Molecules of Biological Interest, Düsseldorf Germany, 06/2001 23. “DNA Building Blocks in the Gas-Phase”, Molecular & Ionic Clusters, Gordon conference, Ventura, CA, 01/2002 24. “Measuring Fast Conformational Dynamics of Freely Diffusing Single Molecules”, Spectroscopy of Biomolecular Dynamics, Kloster Banz, Germany, 03/2004 25. “Fast Conformational Dynamics of Freely Diffusing DNA-Hairpin Using FCS”, 227th ACS National Meeting, Anaheim, CA, USA, 03/2004 26. “Fast Conformational Dynamics of Freely Diffusing DNA-Hairpin Extracted Using FCS”, 228th ACS National Meeting, Philadelphia, PA, USA, 08/2004 27. “Fast Conformational Dynamics of Freely Diffusing Single DNA-Hairpin Extracted From sp-FRET Histogram Shape Analysis”, 49th Biophysical Society, Long-Beach, CA, USA, 02/2005 28. “Fast Conformational Dynamics of Freely Diffusing Single DNA-Hairpin Extracted From sp-FRET Histogram Shape Analysis”, 229TH ACS National Meeting, San Diego CA USA, 03/2005 29. “Measuring Fast Conformational Dynamics of Freely Diffusing Single Molecules”, 5th EBSA international Biophysics congress, Montpellier, France, 09/2005 30. “Measuring fast Conformational Dynamics of Freely Diffusing Single Molecules by FRET-Histogram Shape Analysis”, Frontiers in Chemical Biology, Single Molecules, Cambridge, UK, 03/2006 31. “Studying the Structural Dynamics of Autonomous Bipedal DNA Motors with Single- Molecule Fluorescence Spectroscopy”, The 18th International Conference on DNA Computing and Molecular Programming, Aarhus, Denmark, 08/2012 32. “Fast and Efficient DNA-Based Molecular Motors Assisted by Microfluidics and Single-Molecule Fluorescence”, Single Molecule Approaches to Biology, Gordon Research Conference, Lucca, Italy, 07/2014
(c) Seminar Presentations at Universities and Institutions 33. Physical Chemistry seminar, Heinrich Heine university, Dusseldorf, Germany, 08/2001, (Two Talks) 34. Biophysical seminar, Department of Chemistry-Biochemistry, UCLA, Los-Angeles, US, 06/2007, (Talk) Dr. Eyal Nir page 9
35. Physical Chemistry seminar, Tel-Aviv University, Tel-Aviv, Israel, 01/2012, (Talk) 36. Departmental seminar, The German Cancer Research Center, DKFZ, Heidelberg, Germany, 09/2012, (Talk) 37. Biophysical seminar, Bar-Ilan University, Ramat-Gan, Israel, 11/2012, (Talk) 38. Nano-Seminar arranged by the group of Dr. Tim Liedl, Ludwig Maximilians University, Munich, Germany, 02/2013, (Talk) 39. DNA-Nanotechnology consortium, Technique University of Munich, Munich, Germany, 02/2013, (Talk) 40. Nano-Seminar arranged by the groups of Dr. Turberfield, Dr. Kapanidis and Dr. Louis, Oxford University, Oxford, UK, 02/2013, (Talk) 41. Departmental (Chemistry) seminar, Technion, Israel, 04/2013, (Talk) 42. Physical Chemistry seminar, Ludwig Maximilians University of Munich, Munich, Germany, 12/2013, (Talk)
(d) Additional Participation in Conferences 43. Protein Folding Dynamics, Gordon conference, Ventura, CA, 01/2004 44. The 74th Meeting of the Israeli Chemical Society, Tel-Aviv, Israel, 02/2009 45. Symposium in Chemical Physics, Biological Soft Matter, Tel-Aviv, Israel, 06/2009 46. NanoIsrael 2010, Tel-Aviv, Israel, 2010 47. Nucleosome positioning, chromatin structure and evolution, University of Haifa, Israel, 05/2012 48. Protein folding: Moving beyond simple model systems, Weizmann Institute, Rehovot, Israel, 05/2012 49. NanoIsrael 2012, Tel-Aviv, Israel, 2012
(e) Students Presentations 50. Masoud, R.; Tsukanov, R.; Tomov, T. E.; Plavner, N.; Liber, M; Nir, E., “Designing, Measuring and Improving DNA-Motors using Single-Molecule Fluorescence Spectroscopy”, NanoIsrael, 2010 51. Masoud, R.; Tsukanov, R.; Tomov, T. E.; Plavner, N.; Liber, M.; Nir, E., “Studying Bipedal DNA Motors with Single-Molecule Fluorescence Spectroscopy”, Israel Biophysical Society, 2011 52. Tomov, T. E.; Tsukanov, R.; Masoud, R.; Liber, M.; Plavner, N.; Nir, E., “Disentangling Subpopulations in Single-Molecule FRET and ALEX Experiments with Photon Distribution Analysis”, Israel Biophysical Society, 2011 Dr. Eyal Nir page 10
53. Tsukanov, R.; Tomov, T. E.; Masoud, R.; Drory, H.; Plavner, N.; Liber, M; Nir, E., “Studying DNA hairpin Using Single Molecule Spectroscopy”, Israel Biophysical Society, 2011 54. Plavner, N.; and Nir, E., “Nucleosome Structural Dynamics Addressed by Single Molecule Spectroscopy”, Israel Society for Experimental Biology (ILANIT), 2011 55. Plavner, N.; Tsukanov, R.; Tomov, T. E.; Masoud, R.; Nir, E., "Nucleosome Core Particle Investigated Using Single-Molecule Fluorescence Spectroscopy", Nucleosome positioning, chromatin structure and evolution, Haifa, Israel, 2012 56. Plavner, N.; Tsukanov, R.; Tomov, T. E.; Masoud, R.; Nir, E., "Nucleosome Core Particle Investigated Using Single-Molecule Fluorescence Spectroscopy", Chromatin: from structure to epigenetics, IGBMC, Strasbourg, France, 2012 57. Plavner, N.; Tsukanov, R.; Tomov, T. E.; Masoud, R.; Nir, E., "Nucleosome Core Particle Investigated Using Single-Molecule Fluorescence Spectroscopy", Departmental Talk, DKFZ, Heidelberg, Germany, 2012 58. Plavner, N.; Tsukanov, R.; Tomov, T. E.; Masoud, R.; Nir, E., "Nucleosome Core Particle Investigated Using Single-Molecule Fluorescence Spectroscopy", Protein folding: Moving beyond simple model systems, Weizmann Institute, Rehovot, Israel, 2012 59. Tsukanov, R.; Tomov, T. E.; Masoud, R.; Drory, H.; Plavner, N.; Liber, M.; Nir, E. “DNA Origami as a template for structural dynamics investigation of biomolecules; DNA hairpin”, Protein folding: Moving beyond simple model systems, Weizmann Institute, Rehovot, Israel, 2012 60. Tsukanov, R.; Nir, E., “Measuring DNA Hairpin Dynamics from Several Microsecond to Several Seconds Using Single Molecule Fluorescence Spectroscopy”, NanoIsrael, 2012 61. Tomov, T. E.; Tsukanov, R.; Liber, M.; Plavner, N.; Nir, E., “Towards a reliable and Useful DNA-Motors and Devices: The Single-Molecule Fluorescence Strategy”, NanoIsrael, 2012 62. Masoud, R.; Tsukanov, R.; Tomov, T. E.; Plavner, N.; Liber, M.; Nir, E., “Studying Bipedal DNA Motors with Single-Molecule Fluorescence Spectroscopy”, NanoIsrael, 2012 63. Tsukanov, R.; Tomov, T. E.; Masoud, R.; Drory, H.; Plavner, N.; Liber, M.; Nir, E., “DNA Origami as a Template for Structural Dynamics Investigation of Biomolecules; DNA Hairpin Dynamics”, The 18th International Conference on DNA Computing and Molecular Programming, Aarhus, Denmark, 2012 64. Tomov, T. E.; Tsukanov, R.; Liber, M.; Plavner, N.; Nir, E., “Towards a Reliable and Useful DNA-Motors and Devices: The Single-Molecule Fluorescence Strategy”, The Dr. Eyal Nir page 11
18th International Conference on DNA Computing and Molecular Programming, Aarhus, Denmark, 2012 65. Tsukanov, R.; Tomov, T. E.; Masoud, R.; Drory, H.; Plavner, N.; Liber, M.; Nir, E. “DNA Origami as a Template for Structural Dynamics Investigation of Biomolecules; DNA Hairpin Dynamics”, The 18th Single Molecule workshop, PicoQuant, Berlin, Germany, 2012 66. Tomov, T. E.; Tsukanov, R.; Liber, M.; Plavner, N.; Nir, E., “Towards a Reliable and Useful DNA-Motors and Devices: The Single-Molecule Fluorescence Strategy”, The 18th Single Molecule workshop, PicoQuant, Berlin, Germany, 2012 67. Tomov, T. E.; Tsukanov, R.; Liber, M.; Berger, Y.; Plavner, N.; Nir, E., “Non- Autonomous Motor operate inside microfluidics device”, The 10th Annual FNANO conference (the most important conference in the field), Utah, US, 2013 – INVITED SPEAKER 68. Tomov, T.E.; Tsukanov, R.; Masoud, R.; Liber, M.; Plavner, N.; Nir, E., “Disentangling Subpopulations in Single-Molecule FRET and ALEX Experiments with Photon Distribution Analysis”, Single Molecule Biophysics Seminar Nave Ilan, Israel, 2013 69. Liber, M.; Tsukanov, R.; Tomov, T.E.; Masoud, R.;Plavner, N.; Nir, E., “The Third dimension in DNA Origami Structures – Stacking Origami”, Single Molecule Biophysics Seminar Nave Ilan, Israel, 2013 70. Tsukanov, R.; Tomov, T.E; Masoud, R.; Drory, H.; Plavner, N.; Liber, N.; Nir, E., “Detailed Study of DNA Hairpin Dynamics using Single-Molecule Fluorescence Assisted by DNA Origami”, Single Molecule Biophysics Seminar Nave Ilan, Israel, 2013 71. Plavner, N.; Tsukanov, R.; Tomov, T. E.; Masoud, R.; Nir, E., "Nucleosome Core Particle Investigated Using Single-Molecule Fluorescence Spectroscopy", Single Molecule Biophysics Seminar Nave Ilan, Israel, 2013 72. Plavner, N.; Tsukanov, R.; Tomov, T. E.; Masoud, R.; Nir, E., "Nucleosome Core Particle Investigated Using Single-Molecule Fluorescence Spectroscopy", The 5th Annual Meeting of the Israel Biophysical Society, Israel, 2013 73. Tomov, T.E.; Tsukanov, R.; Liber, M.; Plavner, N.; Nir, E., “Towards a Reliable and Useful DNA-Motors and Devices: The Single-Molecule Fluorescence Strategy”, The 5th Annual Meeting of the Israel Biophysical Society, Israel, 2013 74. Tsukanov, R.; Tomov, T.E.; Berger, Y.; Liber, M.; Nir, E., “Conformational Dynamics of DNA Hairpins at Milliseconds Resolution Obtained from Analysis of Single- Molecule FRET Histograms”, The 5th Annual Meeting of the Israel Biophysical Society, Israel, 2013 Dr. Eyal Nir page 12
75. Liber, M.; Tsukanov, R.; Tomov, T.E.; Masoud, R.; Plavner, N.; Nir, E., “The Third dimension in DNA Origami Structures – Stacking Origami”, The 5th Annual Meeting of the Israel Biophysical Society, Israel, 2013 - Best Poster Award 76. Tomov, T.E.; Tsukanov, R.; Liber, M.; and Nir, E., “Towards a Reliable and Useful DNA-Motors and Devices: The Single-Molecule Fluorescence Strategy”, The 7th Schulich Graduate Students Symposium, Technion, Haifa, Israel, 2013 77. Tomov, T.E.; Tsukanov, R.; Glick, Y.; Liber, M.; Berger, Y.; Gerber, D.; Nir, E., “Two Steps Forward for a Fast and Efficient Non-Autonomous DNA Motor”, NanoIsrael- 2014, Tel Aviv, Israel, 2014 78. Tomov, T.E.; Tsukanov, R.; Glick, Y.; Liber, M.; Berger, Y.; Gerber, D.; Nir, E., “Toward Fast and Efficient DNA Based Molecular Machines: using Single-Molecule Fluorescence and Microfluidics”, Single Molecule Approaches to Biology, Gordon Research Conference, Lucca (Barga), Italy, 2014 79. Tomov, T.E.; Tsukanov, R.; Liber, M.; Glick, Y.; Berger, Y.; Gerber, D.; Nir, E., “Toward Fast and Efficient DNA-Based Molecular Machines: using Single-Molecule Fluorescence and Microfluidics”, 20th International Workshop on Single Molecule Spectroscopy and Ultra-Sensitive Analysis in the Life Sciences, PicoQuant, Berlin, Germany, 2014 80. Tsukanov, R.; Tomov, T.E.; Berger, Y.; Liber, M.; Nir, E., “DNA Hairpin Dynamics Studied by Single Molecule Fluorescence Spectroscopy”, Physical and Theoretical Chemistry Department, Technische Universität Braunschweig, Germany, 2014 - Departmental Seminar Talk 81. Tsukanov, R.; Tomov, T.E.; Berger, Y.; Liber, M.; Nir, E., “Conformational Dynamics of DNA Hairpins at Milliseconds Resolution Obtained from Analysis of Single- Molecule FRET Histograms”, NanoIsrael-2014, Tel Aviv, Israel, 2014 82. Tsukanov, R.; Tomov, T.E. ; Berger, Y. ; Liber, M. ; and Nir, E., “Conformational Dynamics of DNA Hairpins at Milliseconds Resolution Obtained from Analysis of Single-Molecule FRET Histograms”, Ilse Katz Institute Day 2012, Ben Gurion University of the Negev, Israel, 2014 83. Tsukanov, R.; Tomov, T.E.; Berger, Y.; Liber, M.; Nir, E., “Conformational Dynamics of DNA Hairpins at Milliseconds Resolution Obtained from Analysis of Single- Molecule FRET Histograms”, The 20th International Workshop on Single Molecule Spectroscopy and Ultra-Sensitive Analysis in the Life Sciences, PicoQuant, Berlin, Germany, 2014 - Talk 84. Berger, Y.; Tsukanov, R.; Tomov, T.E.; Liber, M.; Nir, E., "Multi Legged DNA- Origami-Based Motors", NanoIsrael-2014, Tel Aviv, Israel, 3/2014 Dr. Eyal Nir page 13
85. Berger, Y.; Tsukanov, R.; Tomov, T.E.; Liber, M.; Nir, E., "Multi Legged DNA- Origami-Based Motors”, Ilse Katz Institute Day, Ben Gurion University of the Negev, Israel, 2014 86. Berger, Y.; Tsukanov, R.; Tomov, T.E.; Liber, M.; Nir, E., "Multi Legged DNA- Origami-Based Motors”, The 20th International Workshop on Single Molecule Spectroscopy and Ultra-Sensitive Analysis in the Life Sciences, PicoQuant, Berlin, Germany, 2014 87. Liber, M.; Tsukanov, R.; Tomov, T.E.; Masoud, R.; Nir, E., “The Third dimension in DNA Origami Structures – Stacking Origami”, NanoIsrael-2014, Tel Aviv, Israel, 2014 88. Liber, M.; Tomov, T.E.; Tsukanov, R.; Berger, Y.; Nir, E., "Development of High- Ordered DNA Origami Structures for DNA Dynamic Machines and Devices”, The Israel Vacuum Society Student Conference, Israel, 2014 - Talk
Grants 1. Focal Technology Area (FTA) grant – “Bio-inspired Nano-carriers for Sub-Cellular Targeted Therapeutics” – together with a group of 10 other PI’s, 680K NIS/Year (each PI) 2012-2016. 2. Israel Science Foundation: “Combining DNA-nanotechnology, microfluidics and single-molecule fluorescence techniques to develop DNA-nanodevice capable of manipulating biomolecules” 230K NIS/Year for 2013-2017.
Synopsis
Many of the most important chemical process taking places in living systems involve large macromolecules such as DNA, RNA, and proteins. Some of these macromolecules have only structural roles, whereas others function by consuming chemical energy and performing chemical (enzymatic) or mechanical work. The latter, therefore, can be perceived as molecular machines. These machines often operate with chemical yields and efficiencies unmatched by artificially made macro and molecular machines. Inspired by biological machines my group seek to rationally design efficient and reliable molecular machines able to perform useful chemical and mechanical work. To design artificial machines and to better understand living systems, it is essential to understand the structure, dynamics, interactions, and functions of complex macromolecules and biological molecular machines on a molecular level. Our group develops and uses a variety of single- molecule fluorescence (SMF) tools exactly for these purposes. As detailed below, in recent years we have combined microfluidics technology with SMF, we have developed photon statistical analysis procedures to identify dynamics and stoichiometry fluctuations, we have verified methods to extract dynamics from the shape of Förster resonance energy transfer (FRET) histograms, and we are involved in the development of coarse-grained simulations of DNA dynamics. We use these and other SMF tools for the development of robust and efficiently operating bipedal motors made of DNA, for studying dynamics of DNA hairpin model systems in unprecedented detail and resolution, for detailed study of the mechanisms of nucleosome core particle association and dissociation, and Dr. Eyal Nir page 14
for the study and characterization of polysaccharide/DNA nano-carriers that may be useful for drug delivery.
1. DNA-based Molecular Machines: The manufacturing of efficiently functioning machines is a major challenge of modern technology and science. DNA nanotechnology is probably the most promising immediate path toward realization of this goal on the molecular level. A major goal of my group is to develop reliable and robust DNA devices that are able to manipulate and maneuver other molecules. Our immediate goal is to design a bipedal motor that is capable of conducting hundreds of sequential steps, striding over several micrometers, and carrying cargo. We have recently achieved three major steps toward realization of this model system: First, we recognized a major problem with the design of previous ‘fuels’ for these types of motors (i.e., the DNA strands that provide chemical energy to the motor). We solved this problem by rationally designing fuels containing hairpin structure that dictate hierarchal interaction with the motor; use of these fuels increases the operational yield by 74 fold (Journal of American Chemical Society, 2012, Ref. #25). Second, we demonstrated that by operating the motor inside a microfluidics device (in collaboration with the group of Dr. Doron Gerber from Bar-Ilan University) the motor successfully strides over 400 nm, conducting 72 sequential chemical reactions with 50% operational yield. To the best of our knowledge, this is the highest number of operations ever demonstrated for an artificial molecular machine. Third, we demonstrated that track that are several micrometers long can be prepared by polymerizing DNA origami tiles and that the bipedal motor can cross between the tiles without dissociating, opening the door for motors that are able to stride over several micrometers (Small, 2014, Ref. #29). Encouraged by these successes we envision a variety of molecular machines that will be able to pull, push, twist, bend, and maneuver other molecules, operated inside the microfluidics device and controlled by a computer with monitoring by SMF. We expect that these devices will be able to carry out syntheses of molecules that are otherwise impossible. In another example, we reconstructed a previously published bipedal motor that was design to operate autonomously. As a test case for the capabilities of the SMF method we showed that it is possible to monitor the various steps of the motor assembly and operation, as was demonstrated before using traditional gel electrophoresis technique. Moreover, with SMF, we were able to monitor the motor assembly and operation kinetics, impossible using gel electrophoresis and analysis of the data revealed problems with the design and the preparation strategy (ACS Nano, 2012, Ref #24). Based on this conclusion, we developed the microfluidics strategy that turn out to be very successful. These projects are part of a broader effort to construct a DNA puller, an effort funded by an ISF grant (2013-2016).
2. DNA Dynamics and Development of Coarse-Grained Simulations: Another major effort in our group is the study of DNA dynamics. Our goal is to understand the structure, dynamics, and interactions of simple and complex DNA (and eventually RNA) systems. These studies will further our understanding of interactions between DNA and proteins and DNA and RNA folding and will rational design process in DNA nanotechnology. Using single-molecule FRET probability distribution analysis (PDA, see below) we have measured and analyzed the opening and closing rates of two-state DNA hairpins with different loop and stem sequences and lengths at resolution impossible using other methods (part of this work was published in two papers in Journal of Physical Chemistry B, 2013, Ref #26 and Ref #27). We were able to make general conclusions about the influence of the hairpin loop and stem sequences and the buffer ionic strength on the hairpin dynamics and were able to characterize the transition state. To gain better and more detailed understanding of the hairpin dynamics we established collaborations with the groups of Prof. Louis and Prof. Doye from Oxford University, experts in coarse-grained simulations of DNA. Using our data, these groups have further developed the simulation’s force field to include base stacking and the influence of counter ions. Using very minimal parameterization the simulated data agree very well with the experimental results. Importantly this research has confirmed the non- trivial influence of the hairpin stems on the closing rates. These very promising results encouraged our groups to extend the collaboration to more complex systems. For example, we are testing the optimal step size for our relatively complex bipedal motor, and initial results show good agreement Dr. Eyal Nir page 15
between experiments and simulations. We envision that the close collaboration between our lab and the Oxford theoretical groups will result in a coarse-grained tool that will enable in silico rational design of DNA devices and machines and accurate prediction of the structures adopted by RNA and DNA molecules. Although not proposed originally, the development of the coarse-grained tool will have direct impact on the design of the device proposed in the ISF grant (2013-2016).
3. Dynamics of Nucleosome Core Particles: The stability of the nucleosome core particle (NCP, consisting of eight histone proteins and double-stranded DNA) depends on DNA sequence and is believed to play a major role in the regulation of gene expression. To understand the mechanisms that dictate NCP stability, we are studying NCP dissociation and association mechanisms and determining dependence on ionic strength and DNA sequence. We observed a non-monotonous dependency of NCP stability on NaCl concentration. We suggest that this rather unexpected behavior is a result of delicate interplay between repulsive and attractive forces between positively charged histones themselves and between the histones and the negatively charged DNA. Interestingly, we found that under some conditions the NCP is least stable at near physiological ionic strength and postulate that this may indicate that changes in ionic strength in the cell nucleus trigger the release of histone proteins to facilitate chromosome separation and replication during cell division.
4. Characterization of Nano-Carriers for Drug Delivery: As part of our participation in the Focal Technology Areas (FTA) grant, a collaboration among eleven groups (2012-2016), we are investigating and characterizing the size, content, stoichiometry, and stability of polysaccharide/DNA nano-carrier under different conditions. We hypothesize that these carriers will be useful for sub-cellular targeting and drug delivery. We adapted the alternating laser excitation (ALEX) and the total internal reflection fluorescence (TIRF) techniques specifically for these purposes. The polysaccharide/DNA stoichiometry and the size of the particle have been characterized, and the nano-carrier stability and propensity for aggregation at different concentrations and in the presence of different ions were determined.
5. Methods Development: My group is involved in developing the SMF experimental toolkit. Below are some of our recent accomplishments.
Two Channel Density Estimator: In a diffusion-based SMF experiment, molecules diffuse in an out of the confocal spot producing bursts of photons, each corresponding to an individual molecule. We have developed a novel approach to identify fluctuations in FRET and in dye stoichiometry. Our method is based on statistical analysis of the distribution of the times of photon arrival inside a burst. We call this tool the two channel density estimator (2CDE, Biophysical Journal, 2012, Ref. #23). Using this tool, we revealed the underling two-state structure of a hairpin molecule that was concealed in dynamic heterogeneity, identified minor intact bipedal motor subpopulations otherwise hidden in the ensemble, and filtered for a minor population of doubly-labeled double-stranded DNA from the much larger population of single-stranded (and singly labeled) DNA. We are aware of at least four groups that are already using the 2CDE method to identify, characterize, and filter subpopulations in their data (from publications and private communications).
Probability Distribution Analysis: PDA is a single-molecule method for extracting transition rates between states by analyzing the shape of FRET histograms. The method was developed by me (during my post-doctorate studies and during my Tenure-Track,(Statistic & Probability Letters, 2010, Ref. #22) and by other researcher groups before, but had never been experimentally validated. By comparison of data collected on sets of two-state hairpin molecules and comparison to results acquired from surface immobilized hairpins and hairpins attached to origami we demonstrated the validity of PDA and provided a simple but robust algorithm to implement PDA (Journal of Physical Chemistry B, 2013, Ref #27). Furthermore, we showed very good agreement between rates acquired from immobilization-based and diffusion-based measurements revalidating Dr. Eyal Nir page 16
these two methods. As our data indicated that the origami had no influence on hairpin dynamics, origami provides a platform on which biomolecules can be reliably investigated.
Combining Microfluidics Technology with SMF: In collaboration with the group of Dr. Gerber, we successfully combined a microfluidic device containing computer-controlled pneumatic valves (Quake type valves that are imbedded inside the device) with single-molecule TIRF setup. This combination (novel, to the best of our knowledge) enabled computer-controlled sequential introduction of 72 fuel and antifuel strands to allow a bipedal motor to stride over 400 nm, while the motor was continually monitored using SMF. This record-high operational yield was achieved because the motor was always attached to the surface and only the solutions containing the strands were replaced, similar to the principle of solid-phase synthesis, a method known for its high chemical yield. The fast replacement of the solution enabled by the microfluidics device allow us to make kinetic measurements with 0.1 s resolution, more than three orders of magnitude faster than possible with diffusion-based SMF. We used this technique to study the dynamics of motor leg- placing and leg-lifting reactions and were able to determine that the minimum time it takes for the motor to step forward is around 1 s. We anticipate that the combination of these two technologies will enable assembly, operation, and characterization of dynamic molecular devices and structures impossible otherwise. The usage of microfluidics was proposed in an ISF grant (2013-2016).
Single-Molecule Fluorescence as a Tool of Choice for the Development of DNA Nanotechnology and Molecular Machines: Because of our contribution to the field of DNA nanotechnology, we were asked by the editors of a recent special issue on DNA nanotechnology in Accounts of Chemical Research (2014, Ref #28) to review the contribution of SMF to the field. We presented several examples (mainly ours) that demonstrate how SMF tools can be utilized for the development of DNA machines and devices and for structural and dynamic investigation of other biomolecules. I strongly believe that our group will continue the successful development of DNA molecular machines, including machines that may perform useful work impossible otherwise.
Present Academic Activities
Manuscript in Preparation 1. Tomov et al., Fast and Efficient DNA Motors Operate using Microfluidics Device and Monitored with Single-Molecule Fluorescence. 2. Plavner et al., On the Association and Dissociation Mechanisms of Nucleosome Core Particles. 3. Sen et al., Characterizing the Size, Content, Stability and Structure of Polysaccharide/DNA Complex using Single-Molecule Fluorescence. 4. In collaboration with the groups of Prof. Louis and Prof. Doye. A paper about optimization of coarse-grained simulations using our hairpin data.