UNIVERSITY OF CALIFORNIA, SAN DIEGO Enhancing Natural Products Structural Dereplication and Elucidation with Deep Learning Based Nuclear Magnetic Resonance Techniques A dissertation submitted in partial satisfaction of the requirements for the degree Doctor of Philosophy in NanoEngineering by Chen Zhang Committee in charge: William H. Gerwick, Chair Garrison W. Cottrell, Co-Chair Gaurav Arya Seth M. Cohen Chambers C. Hughes Preston B. Landon Liangfang Zhang 2017 Copyright Chen Zhang, 2017 All rights reserved The Dissertation of Chen Zhang is approved, and it is acceptable in quality and form for publication on microfilm and electronically: ____________________________________________ ____________________________________________ ____________________________________________ ____________________________________________ ____________________________________________ ____________________________________________ Co-Chair ____________________________________________ Chair University of California, San Diego 2017 iii DEDICATION I dedicate my dissertation work to my family and many friends. A special feeling of gratitude to my beloved parents, Jianping Zhao and Xiaojing Zhang whose words of encouragement and push for tenacity ring in my ears. My cousin Min Zhang has never left my side and is very special. I also dedicate this dissertation to my mentors who have shown me fascinating views of the world throughout the process, and those who have walked me through the valley of the shadow of frustration. I will always appreciate all they have done, especially Bill Gerwick, Gary Cottrell and Preston Landon for showing me the gate to new frontiers, Sylvia Evans, Pieter Dorrestein, Shu Chien, Liangfang Zhang, and Gaurav Arya for their great encouragement, and Wood Lee and Yezifeng for initially showing me the value of freedom, and continuously answering my questions regarding social sciences, humanities, literature, and arts. I dedicate this work and give special thanks to my wonderful friends, especially Niu Du, Sima Yazdani, Yerlan Idelbayev, Jie Min, Zheng Long, Mike Shao, Bettina Lehman, Sheri Harris and Han Huang for not giving me up throughout the entire doctorate program. All of you have been my best cheerleaders. iv EPIGRAPH “So doth the woodbine, the sweet honeysuckle Gently entwist; the female ivy so Enrings the barky fingers of the elm. O, how I love thee! how I dote on thee!” --“A Midsummer Night’s Dream” ACT IV, sc. i., By W. Shakespeare v TABLE OF CONTENTS SIGNATURE PAGE ............................................................................................... iii DEDICATION ........................................................................................................ iv EPIGRAPH ............................................................................................................... v TABLE OF CONTENTS ........................................................................................ vi LIST OF ABBREVIATIONS ................................................................................ xii LIST OF FIGURES ............................................................................................... xiv LIST OF TABLES ............................................................................................... xvii ACKNOWLEDGEMENTS ................................................................................ xviii VITA ...................................................................................................................... xxi ABSTRACT OF THE DISSERTATION ............................................................ xxiii CHAPTER 1 ............................................................................................................. 1 INTRODUCTION .................................................................................................... 1 1.1 Uses of Natural Products in the History ......................................................... 1 1.2 Acquiring Natural Products in the History (Isolation Methods) .................. 11 1.3 Potentials of Natural Products Industrialization ........................................... 16 1.4 Historical Investigations into Marine Natural Products ............................... 21 vi 1.5 Techniques that Enhance Marine Natural Products Discovery .................... 34 1.5.1. Single-Scan 2D NMR Spectroscopy .................................................... 36 1.5.2. 2D Hadamard Spectroscopy ................................................................. 37 1.5.3. Sparse Sampling NMR Spectroscopy .................................................. 37 1.5.4. 2D NMR Spectra Analysis via Machine Learning ............................... 38 1.5.5. Absolute Structural Assignment via ECCD ......................................... 39 1.6 Dissertation Contents .................................................................................... 41 1.7 Chapter 1 References .................................................................................... 43 CHAPTER 2 ........................................................................................................... 59 ISOLATION AND STRUCTURAL ELUCIDATION OF LAUCYSTEINAMIDE A, A HYBRID PKS/NRPS METABOLITE FROM A SAIPAN CYANOBACTERIUM, CF. CALDORA PENICILLATA ......................................... 59 2.0. Abstract ........................................................................................................ 59 2.1 Introduction .................................................................................................. 59 2.2. Results and Discussion ................................................................................ 62 2.2.1. Stereochemistry .................................................................................... 69 2.2.2. Bioactivity ............................................................................................ 71 2.2.3. Biosynthetic Considerations ................................................................. 72 2.3 Conclusion .................................................................................................... 73 2.4 Experimental Methods .................................................................................. 73 vii 2.4.1. General Experimental Procedures ........................................................ 73 2.4.2. Sample Material .................................................................................... 74 2.4.3. Extraction and Isolation ........................................................................ 75 2.4.4. Molecular Networking .......................................................................... 77 2.4.5. Biological Testing ................................................................................ 77 2.5 Chapter 2 Acknowledgements ...................................................................... 78 2.6 Chapter 2 Appendix ...................................................................................... 79 2.7 Chapter 2 References .................................................................................... 93 CHAPTER 3 ........................................................................................................... 96 DEVELOPING SMALL MOLECULE ACCURATE RECOGNITION TECHNOLOGY (SMART), A DEEP LEARNING AND FAST 2D NMR BASED DIGITAL FRONTIER TO ENHANCE NATURAL PRODUCTS DISCOVERY ..... 96 3.0. Abstract ........................................................................................................ 96 3.1 Introduction .................................................................................................. 97 3.2 Results and Discussion ............................................................................... 102 3.2.1. The SMART Prototype ...................................................................... 102 3.2.2. Network Training and Results ............................................................ 110 3.2.3. Related Work ...................................................................................... 114 3.2.4. SMART recognition of noisy HSQC spectra ..................................... 118 3.2.5. SMART characterization of Viequeamides of NRPS origin .............. 121 viii 3.3 Conclusion .................................................................................................. 122 3.4 Experimental Methods ................................................................................ 123 3.4.1. Training Set Collection and Processing ............................................. 123 3.4.2. NUS 2D NMR Data Generation ......................................................... 125 3.4.3. The Deep Siamese Network ............................................................... 126 3.4.4. Loss Function ..................................................................................... 127 3.4.5. Training Details of the Siamese Network .......................................... 128 3.4.6. Validation of the Model on “Novel” Categories ................................ 133 3.4.7. Tanimoto Score Calculation ............................................................... 134 3.4.8. Recognition of noisy HSQC spectra ................................................... 134 3.5 Chapter 3 Acknowledgements .................................................................... 136 3.6 Chapter 3 Appendix .................................................................................... 137 3.7 Chapter 3 References .................................................................................. 177