Springer Series in Biophysics 13 Nils G. Walter (cid:129) Sarah A. Woodson (cid:129) Robert T. Batey Editors Non-Protein Coding RNAs 123 Editors Dr. Nils G. Walter Dr. Sarah A. Woodson Associate Professor of Chemistry Professor of Biophysics Department of Chemistry Department of Biophysics University of Michigan Johns Hopkins University 930 N. University 3400 North Charles Street Ann Arbor, MI 48109-1055 Baltimore, MD 21218 USA USA Dr. Robert T. Batey Associate Professor of Chemistry and Biochemistry Department of Chemistry and Biochemistry University of Colorado at Boulder Box 215, Boulder, CO 80309-0215 USA ISSN 0932-2353 ISBN 978-3-540-70833-9 e-ISBN 978-3-540-70840-7 Library of Congress Control Number: 2008931054 © 2009 Springer-Verlag Berlin Heidelberg This work is subject to copyright. 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Cover design: WMXDesign GmbH, Heidelberg, Germany Printed on acid-free paper 9 8 7 6 5 4 3 2 1 springer.com Preface The 2006 Nobel Prize in Physiology or Medicine was awarded to the discoverers of RNA interference, Andrew Fire and Craig Mello. This prize, which follows “RNA” Nobels for splicing and RNA catalysis, highlights just one class of recently discovered non-protein coding RNAs. Remarkably, non-coding RNAs are thought to outnumber protein coding genes in mammals by perhaps as much as four-fold. In fact, it appears that the complexity of an organism correlates with the fraction of its genome devoted to non-protein coding RNAs. Essential biological processes as diverse as cell differentiation, suppression of infecting viruses and parasitic trans- posons, higher-level organization of eukaryotic chromosomes, and gene expression are found to be largely directed by non-protein coding RNAs. Currently, bioinformatic, high-throughput sequencing, and biochemical approaches are identifying an increasing number of these RNAs. Unfortunately, our ability to characterize the molecular details of these RNAs is significantly lacking. The biophysical study of these RNAs is an emergent field that is unraveling the molecular underpinnings of how RNA fulfills its multitude of roles in sustaining cellular life. The resulting understanding of the physical and chemical processes at the molecular level is critical to our ability to harness RNA for use in biotechnology and human therapy, a prospect that has recently spawned a multi-billion dollar industry. This book assembles chapters from some of the experts in Biophysics of RNA to provide a snapshot of the current status of this dynamic field. While by necessity incomplete, this book aims to survey a number of the better characterized non-pro- tein coding RNAs and the biophysical techniques used to study them. It is written for students and researchers at all levels of accomplishment interested in under- standing how non-protein coding RNAs work and how biophysical and computa- tional approaches can be used to delineate the molecular underpinnings of RNA function. Many topics are approached with the goal of describing how biophysical tools and techniques have been used to address fundamental questions in the biol- ogy of non-protein coding RNAs, rather than a description of RNAs themselves. In this light, we hope that the book will be of particular use to junior scientists seeking to tackle new problems in RNA biology from the vantage of biophysics. v vi Preface Following a foreword featuring a general overview of the lessons from the bio- physical study of RNA, the first three chapters aim to describe how theory, simula- tion, and experimental probing can be used to unveil the thermodynamics and kinetics governing RNA folding and dynamics. Chapters 4–6 are devoted to small self-cleaving ribozymes, as understood through the lens of X-ray crystallography, ensemble and single molecule fluorescence, and chemical probing. Subsequent chapters tackle increasingly complex RNAs and their protein complexes. In partic- ular, Chaps. 7–9 focus upon large ribozymes that use more sophisticated mecha- nisms of catalysis and even recruit proteins to facilitate function in the cellular environment. As genetic regulation appears to be an increasingly important role for non-coding RNAs, Chaps. 10 and 11 concentrate on how X-ray crystallography, NMR spectroscopy, and fluorescence techniques have revealed how riboswitches specifically recognize small molecule metabolites to affect gene expression. Many modern non-protein coding RNAs are assembled into large ribonucleoprotein com- plexes (RNPs) and Chaps. 12–14 yield insights into how these particles are assem- bled to form a functional complex. These large RNP machines are by necessity highly dynamic entities that must adopt a number of conformations, as revealed in studies of the ribosome by cryo-electron microscopy in Chap. 15. Finally, non-cod- ing RNAs often interact with other cellular machineries to enable their function, as discussed in Chaps. 16 and 17. We hope that our selection of topics is both timely and stimulating for the rapidly growing RNA community and beyond. USA Nils G. Walter September 2008 Sarah A. Woodson Robert T. Batey Contents 1 RNA 3D Structural Motifs: Definition, Identification, Annotation, and Database Searching ....................................................... 1 Lorena Nasalean, Jesse Stombaugh, Craig L. Zirbel, and Neocles B. Leontis 2 Theory of RNA Folding: From Hairpins to Ribozymes ......................... 27 D. Thirumalai and Changbong Hyeon 3 Thermodynamics and Kinetics of RNA Unfolding and Refolding ........ 49 Pan T.X. Li and Ignacio Tinoco 4 Ribozyme Catalysis of Phosphodiester Bond Isomerization: The Hammerhead RNA and Its Relatives ............................................... 73 William G. Scott 5 The Small Ribozymes: Common and Diverse Features Observed Through the FRET Lens ........................................................................... 103 Nils G. Walter and Shiamalee Perumal 6 Structure and Mechanism of the glmS Ribozyme ................................... 129 Juliane K. Soukup and Garrett A. Soukup 7 Group I Ribozymes as a Paradigm for RNA Folding and Evolution .... 145 Sarah A. Woodson and Seema Chauhan 8 Group II Introns and Their Protein Collaborators ................................ 167 Amanda Solem, Nora Zingler, Anna Marie Pyle, and Jennifer Li-Pook-Than 9 Understanding the Role of Metal Ions in RNA Folding and Function: Lessons from RNase P, a Ribonucleoprotein Enzyme ............................ 183 Michael E. Harris and Eric L. Christian vii viii Contents 10 Beyond Crystallography: Investigating the Conformational Dynamics of the Purine Riboswitch ....................................................... 215 Colby D. Stoddard and Robert T. Batey 11 Ligand Binding and Conformational Changes in the Purine-Binding Riboswitch Aptamer Domains ..................................... 229 Jonas Noeske, Janina Buck, Jens Wöhnert, and Harald Schwalbe 12 The RNA–Protein Complexes of E. coli Hfq: Form and Function .................................................................................. 249 Taewoo Lee and Andrew L. Feig 13 Assembly of the Human Signal Recognition Particle ........................... 273 Elena Menichelli and Kiyoshi Nagai 14 Forms and Functions of Telomerase RNA ............................................. 285 Kathleen Collins 15 Ribosomal Dynamics: Intrinsic Instability of a Molecular Machine ........................................................................... 303 Haixiao Gao, Jamie LeBarron, and Joachim Frank 16 Biophysical Analyses of IRES RNAs from the Dicistroviridae: Linking Architecture to Function ........................................................... 317 Jeffrey S. Kieft 17 Structure and Gene-Silencing Mechanisms of Small Noncoding RNAs ...................................................................................... 335 Chia-Ying Chu and Tariq M. Rana Index .................................................................................................................. 357 Color Plates ....................................................................................................... 365 Contributors Robert Batey Department of Chemistry and Biochemistry, Campus Box 215, University of Colorado-Boulder, Boulder, CO 80309, USA, [email protected] Janina Buck Institut für Organische Chemie und Chemische Biologie, Zentrum für Biomolekulare Magnetische Resonanz, Johann Wolfgang Goethe-Universität, Max-von-Laue-Strasse 7, N160-314, 60438 Frankfurt am Main, Germany, [email protected] Kathleen Collins Department of Molecular and Cell Biology, University of California at Berkeley, Berkeley, CA 94720-3200, USA, [email protected] Andrew Feig Department of Chemistry, Wayne State University, 5101 Cass Ave., Detroit, MI 48202, USA, [email protected] Joachim Frank Howard Hughes Medical Institute, Department of Biochemistry and Molecular Biophysics and Department of Biological Sciences, Columbia University, 650 West 168th Street, NY 10032, USA, [email protected] Haixiao Gao Wadsworth Center, Empire State Plaza, Albany, NY 12201-0509, USA, [email protected] Michael Harris Center for RNA Molecular Biology, Department of Biochemistry, CWRU - School of Medicine, Cleveland, OH 44106, USA, [email protected] ix x Contributors Jeffrey Kief Department of Biochemistry and Molecular Genetics, Denver School of Medicine, University of Colorado, 12801 East 17th Ave, Rm L18-9110, Aurora, CO 80045, USA, [email protected] Jamie LeBarron Wadsworth Center, Empire State Plaza, Albany, NY 12201-0509, USA, [email protected] Neocles Leontis Department of Chemistry, Bowling Green State University, 141 Overman Hall, Bowling Green, OH 43403, USA, [email protected] Kyoshi Nagai Structural Studies Division, MRC Laboratory of Molecular Biology, Hills Road, Cambridge CB2 2QH, UK, [email protected] Jonas Noeske Institut für Organische Chemie und Chemische Biologie, Zentrum für Biomolekulare Magnetische Resonanz, Johann Wolfgang Goethe-Universität, Max-von-Laue-Strasse 7, N160-314, 60438 Frankfurt am Main, Germany, [email protected] Anna Marie Pyle 266 Whitney Avenue, Room 334A Bass Building, Yale University, New Haven, CT 06511, USA, [email protected] Tariq Rana Department of Biochemistry and Molecular Pharmacology, University of Massachusetts Medical School, Worcester, MA 01605, USA, [email protected] Harald Schwalbe Institut für Organische Chemie und Chemische Biologie, Zentrum für Biomolekulare Magnetische Resonanz, Johann Wolfgang Goethe-Universität, Max-von-Laue-Strasse 7, N160-314, 60438 Frankfurt am Main, Germany, [email protected] William Scott Department of Chemistry, University of California, 1156 High Street, Santa Cruz, CA 95064, USA, [email protected] Garrett Soukup Department of Biomedical Sciences, Creighton University School of Medicine, 2500 California Plaza, Omaha, NE 68178, USA, [email protected] Contributors xi Devarajan (Dave) Thirumalai Department of Chemistry and Biochemistry, University of Maryland, College Park, MD 20742, USA, [email protected] Ignacio Tinoco Department of Chemistry, University of California, Berkeley, CA 94720-1460, USA, [email protected] Olke Uhlenbeck Department of Biochemistry, Molecular Biology and Cell Biology, Hogan 2-100, 2205 Tech Drive, Evanston, IL 60208, USA, [email protected] Nils Walter Department of Chemistry, University of Michigan, Ann Arbor, 930 N. University, MI 48109-1055, USA, [email protected] Jens Wöhnert Institut für Molekulare Biowissenschaften, Zentrum für Biomolekulare Magnetische Resonanz, Johann Wolfgang Goethe-Universität, Max-von-Laue-Strasse 9, N200-2.04, 60438 Frankfurt am Main, Germany, [email protected] Sarah Woodson T.C. Jenkins Department of Biophysics, Johns Hopkins University, 3400 N. Charles St., Baltimore, MD 21218, USA, [email protected]