IMPORTANT INTERACTIONS IN PEPTIDE-BASED TRANSFECTION AGENTS, SUGARS AS CHIRAL SCAFFOLDS, AND MOLECULAR IMPRINTING OF NERVE GASES By FLORENT ALLAIS A DISSERTATION PRESENTED TO THE GRADUATE SCHOOL OF THE UNIVERSITY OF FLORIDA IN PARTIAL FULFILLMENT OF THE REQUIREMENTS FOR THE DEGREE OF DOCTOR OF PHILOSOPHY UNIVERSITY OF FLORIDA 2004 Copyright 2004 by Florent Allais This is dedicated to my parents Nadine and Jacques. This is for Stanislas, especially, who inspires me daily. ACKNOWLEDGMENTS First and foremost I would like to thank my parents and my grandmother Maria. Their love, support, and understanding are a continuous source of strength. They have always continued to support me in whatever endeavor I choose and I will forever be grateful. I would also like to thank my great-grandmother Alberte, who taught me a lot while I was a child; and my grandfather Georges, whom I didn’t have the joy to really know. I am sure they would be really proud to see the individual I became and where I am now. There are other important persons in my life: Stanislas, the greatest man I could have ever found; my family (although some of them did or still behave in a way that disgusts and upsets me); Nathan, (though I haven’t known him for a long time, I know that we will be friends forever); and finally Xavier, who is someone I won’t forget, in spite of huge misunderstandings about life and relationships. Next I would like to thank Pr. Eric Enholm for being a great advisor, but more importantly an excellent teacher. I valued the environment he created in his lab and the many discussions on topics at hand. He made me feel at home when I came to his lab the first time, 5 years ago. Having such a warm and friendly welcome helped me make up my mind when it came to choosing a place for my Ph.D. He cares about his students, and I thank him for his support and guidance over the years. I would also like to thank Dr. Randy Duran, the one who made my dream come true. For years I wanted to come to the USA for my education. Thanks to the REU program, I had the opportunity to spend Summer’99 at the University of Florida and meet amazing and interesting people, and a iv great laboratory. I will always be grateful to the REU program. I would like to thank Dr. Merle Battiste, Dr. Dolbier, Dr. Randy Duran and Dr. Ken Sloan for serving on my committee for their help and sharing of their wisdom over the years; and also for dealing with my terrible French accent (oops, sorry, Freedom accent). Dr. Kenneth Sloan is a great committee member and we should all be so lucky. I would like to thank the Enholm Group members past and present. Jennifer Cottone, Maria Gallagher, Ashwin Bharadwaj, Sophie Klein, Aarti Joshi, Jed Hastings, Emilie Claudel, Rayanne Mohammed, Christophe Blaszykowski, and Sébastian Bareyt made the time spent in the lab a joy, with loud music, constant joking (about Indians, Americans and French), and dance moves. Sometimes we argued, but I don’t know any workplace where everyone gets along 24/7! Special thanks go to Jennifer for training me and taking me under her wing upon arrival. Ryan Martin is a great undergraduate to work with, and I wish him nothing but the best. Despite profound disagreements on specific topics, I enjoyed working with him. He’s a great individual and deserves to succeed. Thanks also go to the many other students in the department who enriched my experience. Thanks go to the “French Mafia”: Chaya Pooput, Christophe Grenier, Thomas Joncheray, Emilie Galand, Daniel Serra, Sophie Bernard, Thomas Cardolaccia, and Florence Courchay. I will miss our parties and delicious lunches/dinners. They were great occasions to cheer us up when home sickness struck. I also want to thank Maria Estrella, Lynn Usher, Ravi and many others for their friendship and valuable discussion. There is chemistry and then there is the home side. After leaving work, it can be difficult to keep work at work. Without these people, I would have had no life outside the work. Thus I would like to thank Chaya Pooput and Sophie Klein for being great v roommates and persons who know what is truly important. All the people named above are not just friends but part of my family and I will miss them. We had many good times and I will miss them. I also want to give special thanks for having been awarded the MacLaughlin Dissertation Fellowship to finance my last semester here at the University of Florida. I really appreciated their recognition of my work and career. I would like to thank the “French faculty”: Pr. Yannick Landais, who taught me rigour and precision; Catherine Grosdemange-Billard, who is an excellent professor and also a great friend who helped (and still helps) me construct my future and career; and the REU coordinators Dr. Anne-Lise Dhimane. Dr. Yves Gnanou. Finally I thank Pr. Janine Cossy for welcoming me into her research group as a postdoctoral fellow. I sincerely hope I’ll fulfill her expectations. vi TABLE OF CONTENTS page ACKNOWLEDGMENTS ................................................................................................. iv LIST OF TABLES............................................................................................................. ix LIST OF FIGURES .............................................................................................................x ABSTRACT...................................................................................................................... xii CHAPTER 1 HISTORICAL BACKGROUND .................................................................................1 History of Free-Radicals...............................................................................................1 Generation of Radicals .................................................................................................2 Free-Radical Cyclizations.............................................................................................4 5-Hexenyl Radical Cyclizations ...................................................................................6 Chiral Templates for Free-Radical Chemistry..............................................................8 DNA Transfection ......................................................................................................12 RGD Analogues..........................................................................................................13 History of Molecular Imprinting ................................................................................19 Methodology and Principles of Molecular Imprinting ...............................................20 Fluorescence and Fluorescent Molecules in MIP.......................................................22 Nerve Gases................................................................................................................24 2 ALKENE-ALDEHYDE CYCLIZATION VIA O-KETYL RADICAL USING LEWIS ACIDS AND SUGARS AS AUXILIARIES ................................................28 Introduction.................................................................................................................28 Preliminary Study .......................................................................................................30 Synthesis of the Chiral Precursors..............................................................................31 Results and Discussion ...............................................................................................34 Conclusion ..................................................................................................................38 3 MOLECULAR IMPRINTING WITH ROMP METHODOLOGY: EFFECTS ON POLYMERIZATION AND IMPRINTED PROPERTIES........................................39 Introduction.................................................................................................................39 Comparison of ROMP versus Radical Induced Polymerization ................................39 vii Synthesis of the MIPs .................................................................................................40 Templates....................................................................................................................40 Radical Polymerization...............................................................................................41 ROMP Polymerization................................................................................................42 Synthesis of the Monomers ........................................................................................45 Analytical Method, Results and Discussion ...............................................................47 Control Experiments...................................................................................................54 Using MIP Technology in the Synthesis of Selective Nerve Gas Detectors..............55 Synthesis of the Sensors .............................................................................................58 Analytical Studies.......................................................................................................63 Conclusion ..................................................................................................................63 4 NEW SYNTHETIC VECTORS FOR DNA TRANSFECTION BASED ON A PROLINE MOIETY...................................................................................................65 Introduction.................................................................................................................65 Synthesis