ANALYSIS OF BIO-MOLECULES FOR SYSTEM LEVEL UNDERSTANDING OF ALZHEIMER’S DISEASE By PRIYA PRADAYANI PANIGRAHI A THESIS SUBMITTED IN FULFILLMENT OF THE REQUIREMENTS FOR THE DEGREE OF DOCTOR OF PHILOSOPHY IN BIOINFORMATICS JAYPEE UNIVERSITY OF INFORMATION TECHNOLOGY WAKNAGHAT OCTOBER, 2014 ACKNOWLEDGEMENTS God gives blessings to us so we can give glory to him. I'm blessed and I thank God for everything in my life. I deem it my privilege and honour to place and record my gratitude and indebtedness to the following without whose support, guidance, and concern I would not have been able to complete my Ph. D thesis. I feel privileged to express my deep sense of reverence and gratitude to my revered mentor, Dr. Tiratha Raj Singh, for his support, immaculate guidance, constructive criticism, constant encouragement and providing requisite facilities to carry on my research which otherwise would have remained incomplete. His nurturing and caring concern has been a stimulus which I will always cherish. I also appreciate his untiring efforts during the entire tenure of my research work and patience during writing of this thesis. I have no words to express my gratitude for everything he has contributed in my Ph. D and without his blessings it was surely impossible for me to finish my work. I thank him from bottom of my heart. I emphatically express my loyal and venerable thanks to Prof. Shiban Kishen Kak (Vice Chancellor, JUIT), Brig. (Retd.) Balbir Singh (Director, JUIT), Prof. T. S.Lamba (Dean, JUIT) and Dr. Y. Medury (Ex-COO, Jaypee Education System) for providing opportunity to pursue a Doctorate Degree, teaching assistantship and advanced lab infrastructure to accomplish this scientific venture of my life. I gratefully acknowledge the help rendered by Prof. R. S. Chauhan (Dean, BT and HOD, Dept. of BT& BI) for his encouragement, timely help and cooperation throughout my research work. It gives me immense pleasure to express my gratitude to Chauhan Sir for his ever smiling disposition coming to my rescue in solving my problems and suggestions which helped me in maintaining my confidence. I also feel indebted to my all time advisor, Mr. Sudipta Ku. Das (Lect. in dept. of Zoology, P.S.College, gaisilat, Odissa), Dr. Ragini Raj Singh (Assist.Prof. Dept. of Physics and Material Science, JUIT), and Dr. Anupriya Kaur (Assist. Prof., Dept. of Humanities and Social Science, JUIT) for their valuable advice, unconditional support and guidance. This document would have remained an infant had it not received its necessary ‘diet’ in the form of comments, suggestions etc. by Dr. C. Rout, Dr. J. Ramana, Dr. Udayabanu M., Dr. Pradeep Kumar Naik, and Dr.Hemant Sood. I am short of words in expressing my thanks to them, for their innovative ideas that shaped this document. I wish to convey my sincere thanks to all the faculty members of Department of Biotechnology and Bioinformatics, for their help and guidance at the various stages of this study. I am also thankful to all the members of technical and non-technical staff of the department for their assistance and valuable contributions. I am fortunate to have friends who have always stood beside me. I extend my heartfelt thanks to Jatin Sir, Varun Sir, Krishna Sir, Bharti Ma’am, Archit, Nikhil, Manika, Ashwani, Tarun, Ankita, Tamanna, Pawan, Dikshita and Deepika for their sustained support and ever needed cooperation. I also thank Asha, Pallavi, Nidhi, Dibyajyoti, and Yamini for their inspiration throughout my life. It is my pleasure to express my gratitude to all research scholars of the Biotechnology & Bioinformatics Department for keeping me blessed with best wishes. It is worthwhile expressing my gratitude to Dr. Ahmed Moussa, Dr. MoacyrComar Junior, and Mr. Ramit Singla who provided me assistance and valuable contributions. The successful culmination of this thesis could not have been possible without Dr. Kamal Raj Pardasani (Professor, Department of Mathematics, Maulana Azad National Institute of Technology, Bhopal, India) and Dr. Brigitte Vannier (Assistant professor of cell biology, University of Poitiers, Poitiers, France). Their critical review and constructive and insightful comments were very helpful in strengthening this Thesis. The purpose of this acknowledgement will be incomplete if I fail to appreciate the moral support and encouragement rendered by my family and my loving husband, Preetam Biswal (he is also one of my greatest advisor). I would like to express my love to my sweet nephew “Aryaveer Nayak” whose innocence refreshed me during my difficult times. The successful culmination of this thesis could not have been possible without the blessings of my loving parents and Inlaws. Without their help I would have been greatly incapacitated. All may not be mentioned, but no one is forgotten. Priya Pradayani Panigrahi Contents CONTENTS List of Tables I List of Figures II-VI Abbreviations VII-X Abstract 1-4 CHAPTER-1 Introduction 5-41 1.1 Introduction 1.2 Dementia 1.3 Diagnosis of dementia 1.4 Causes of dementia 1.5 Types of dementia and their characteristics 1.5.1 Cortical dementias 1.5.2 Sub-cortical dementias 1.6 History of Alzheimer’s disease 1.7 Alzheimer’s disease 1.7.1 Beta-amyloid plaques 1.7.2 Neurofibrillary tangles 1.8 Aβand its association with TAU 1.9 Symptoms and stages of Alzheimer’s disease 1.9.1 Stage 1 (Mild) 1.9.2 Stage 2 (Moderate) 1.9.3 Stage 3 (Severe) 1.10 Types of Alzheimer’s disease 1.10.1 Early onset Alzheimer’s disease (EOAD) 1.10.2 Late onset Alzheimer’s disease (LOAD) 1.10.3 Familial Alzheimer’s disease (FAD) 1.11 Risk factors for Alzheimer’s disease 1.11.1 Non-genetic risk factors 1.11.1.1 Age 1.11.1.2 Education 1.11.1.3 Concomitant health problems 1.11.2 Genetic risk factors 1.11.2.1 Familial Alzheimer’s disease 1.11.2.2 Sporadic Alzheimer’s disease 1.12 Diagnosis of Alzheimer’s disease 1.13 Important facts of Alzheimer’s disease 1.14 Etiological hypothesis for Alzheimer’s disease 1.14.1 Oxidative stress 1.14.2 Inflammation 1.14.3 Cholinergic hypothesis Jaypee University of Information Technology Contents 1.14.4 Cholesterol metabolism 1.15 Treatment of Alzheimer’s disease 1.16 New remedial advances for treatment of Alzheimer’s disease CHAPTER-2 In silico analysis for functional and evolutionary aspects of 42-62 BACE1 and associated Alzheimer’s releted proteins Abstract 2.1 Introduction 2.2 Materials and methods 2.3 Results and discussion 2.4 Conclusion CHAPTER 3 Enrichment analysis for Alzheimer’s disease associated 63-105 pathways and regulatory patterns with aging and other diseases using microarray gene expression and network data Abstract 3.1 Introduction 3.2 Materials and methods 3.2.1 Data 3.2.2 Data pre-processing 3.2.3 Differential gene expression 3.2.4 Clustering of co-expressed genes 3.2.5 Hypergeometric distribution and association of ranked genes 3.2.6 Topological overlap between co-expressed networks and other associated factors 3.2.7 Prioritization of gene candidates with molecular triangulation 3.2.8 Network motif analysis 3.3 Results and discussion 3.3.1 Differential gene expression, clustering of co- expressed genes 3.3.2 Enrichment analysis through co-expressed networks and ranked list of genes 3.3.3 Novel genes, their variants, transcription factors, and miRNA targets 3.3.4 Brain regions and their pathway mapping 3.3.5 Network motifs and their disease associated annotation 3.4 Conclusion Jaypee University of Information Technology Contents CHAPTER 4 A genome wide association study for all key genes identified 106-126 in our earlier research having direct or indirect impact on the metabolic pathways of Alzheimer’s disease: development of a web resource Abstract 4.1 Introduction 4.2 Materials and methods 4.2.1 Database blueprint 4.2.2 Gene and protein screening process 4.2.3 Compilation of genotype data and their LD, and haplotype analysis 4.3 Results and discussion 4.3.1 Findings 4.3.2 Web outline 4.4 Conclusion CHAPTERS 5 Studies to test the binding of THC-Δ9-tetrahydrocannabinol 127-151 and derivatives on acetylcholine binding protein: a virtual screening and molecular docking study Abstract 5.1 Introduction 5.2 Materials and methods 5.2.1 Molecular docking 5.2.1.1 Substrate selection 5.2.1.2 Rigid docking 5.2.1.3 Induced fit docking (IFD) 5.2.2 Toxicity prediction 5.2.3 Prediction of drug like property 5.2.4 Molecular dynamics simulation 5.3 Results and discussion 5.3.1 Selection of potent lead compound 5.3.2 Molecular docking 5.3.3 Molecular dynamics simulation results 5.3.4 TOPKAT and QIKPROP results 5.4 Conclusion Overall conclusions and future prospects 152-157 Bibliography 158-180 Publications and Presentations 181-183 Jaypee University of Information Technology List of tables LIST OF TABLES Table Title Page No. No. 1.1 Proportional pharmacology of presently accepted drugs 32-33 1.2 Proportional clinical pharmacology of presently available drugs 33-34 2.1 Details of Human and Mus musculus BACE1 Sequences 56 BACE1 Interacting proteins identified through protein-protein 2.2 interaction network. Further functional information compiled 58-61 from literature and other annotations Significant number of genes identified through multifaceted integrative approach. 7 different types of analyses applied on all 3.1 74 data types and number of genes identified for all categories indicated against each one respectively Important putative genes identified through manual screening and computational analyses by applying myriad statistical techniques. Abbreviations used are: PPI Gene Set Enrichment Results (PPI), 3.2 80-85 TTGS, miRNA, and Main KEGG File Results (KEGG).