Looking at Ribozymes E-Book

Fabrice Leclerc

• Herausgeber: John Wiley & Sons
• Kategorie: Wissenschaft und neue Technologien
• Sprache: Englisch

Seitenzahl: 256

Veröffentlichungsjahr: 2024

Beschreibung

Behind the neologism "ribozymes" lies a family of fascinating molecules, ribo-enzymes, which have been relatively little studied. These catalytically active RNAs are found in all strata of life, from viruses to the human genome. At the end of the 1970s, the discovery of a catalytic RNA nestled in an intron, followed by another involved in the maturation of transfer RNAs, led to the discovery of new ribozymes and the transition from a strictly "proteocentric" vision, inherited from the dogma of molecular biology, to a more "nucleocentric" one. Since then, a variety of ribozymes have been identified in genomes, where their functions often remain mysterious. Looking at Ribozymes traces the discovery of these molecules and presents a picture of their functional diversity, catalytic mechanisms and distribution within the tree of life.

Leseprobe

Table of Contents

Cover

Table of Contents

Title Page

Copyright Page

Foreword

Preface

Acknowledgments

1 Fundamentals of RNA and Ribozyme Structure

1.1. Sequences and secondary structures

1.2. RNA folding, tertiary structures and 3D

2 Ribozymes and the “Central Dogma” of Molecular Biology

2.1. The discovery of RNA catalysis and the central dogma of molecular biology

2.2. In search of the primordial polymerase

3 The Discovery of Ribozymes

3.1. The discovery of catalysis by autocatalytic introns

3.2. The discovery of RNA catalysis of RNase P

3.3. The first consequences of these discoveries

3.4. The spliceosome, another ribozyme

4 Ribozyme Engineering and the RNA World

4.1. Classification of ribozymes

4.2. Classification of ribozymes according to catalytic mechanism

5 Structures of Ribozymes

5.1. Structures and catalytic mechanisms of ribozymes

5.2. An example of catalysis control: lariat-capping ribozyme

6 Evolution of the Vision of the Catalytic Mechanisms of Ribozymes, the Hammerhead Ribozyme

6.1. Chemistry and catalysis: between general acid/base and metal cations

6.2. Difficulties in interpreting catalysis data

7 The Distribution of Ribozymes in Living Organisms and Molecular Adaptations during Evolution

7.1. Ubiquitous ribozymes

7.2. Selection pressures at work in ribozyme shaping

7.3. Ribozymes in cellular processes: from viroids to eukaryotes

7.4. Very human ribozymes

Conclusion

References

Index

Other titles from ISTE in Biology and Biomedical Engineering

End User License Agreement

List of Tables

Chapter 4

Table 4.1. Classification of ribozymes according to RFam (Gardner et al. 2011;...

Chapter 6

Table 6.1. Strategies for activation of the O2' nucleophile (general base) and...

List of Illustrations

Chapter 1

Figure 1.1. The four nucleotides of RNA form antiparallel double-stranded heli...

Figure 1.2. Nucleotides and base pairs generating antiparallel double-stranded...

Figure 1.3. Illustration of the notion of a dihedral angle and the different d...

Figure 1.4. Illustration of RNA folding on an example based on the lasso-cappe...

Figure 1.5. Symbolizing nucleotides by rectangular triangles allows each edge ...

Figure 1.6. The different representations of a pseudoknot

Figure 1.7. Prokaryotic and eukaryotic loops E

Figure 1.8. The k-turn is an asymmetric inner loop that interrupts the helical...

Figure 1.9. The different modes of interaction between a GNRA loop and its two...

Figure 1.10. An example of an A-minor interaction in the P4–P6 domain of the T...

Figure 1.11. The small ribosomal subunit detects codon–anticodon complementari...

Figure 1.12. The RFam database (Kalvari et al. 2021)6 of the European Bioinfor...

Chapter 2

Figure 2.1. The central “dogma” of molecular biology and its evolution since i...

Figure 2.2. Splicing by group I and II introns and the spliceosome (from W.H. ...

Figure 2.3. The first step of maturation of tRNA by RNase P allows the leader ...

Figure 2.4. The transesterification reaction in group I and II introns and RNa...

Chapter 3

Figure 3.1. Cleavage mechanism by group I introns.

Figure 3.2. Secondary and tertiary structures of RNase P of Thermotoga maritim...

Figure 3.3. Structure of the elongating ribosome of the thermophilic bacterium...

Figure 3.4. The tautomeric forms of a nucleotide are defined as the different ...

Figure 3.5. Some modified tRNA bases.

Figure 3.6. The spliceosome is a molecular machine of extremely complex dynami...

Figure 3.7. Catalytic mechanism of autonucleolytic ribozymes.

Figure 3.8. The symmetrical rolling circle replication mechanism.

Figure 3.9. Crystallographic structures of historical ribozymes (continued).

Figure 3.10. The collection of nucleolytic ribozymes identified since the earl...

Figure 3.11. Summary diagram of the selection method in vitro (Luptak 2016).

Chapter 4

Figure 4.1. Catalytic activities required for viroid replication carried by ho...

Chapter 5

Figure 5.1. a) Structures of minimal (Scott et al. 1995b) and b) complete (Mar...

Figure 5.2. The catalytic mechanism of a hammerhead ribozyme

Figure 5.3. The structure of the hairpin ribozyme and its catalytic mechanism...

Figure 5.4. The glmS riboswitch is also a ribozyme and uses glucosamine-6-phos...

Figure 5.5. Genetic environment, function and structure of the LC ribozyme

Figure 5.6. The process of “homing” in Didymium iridis occurs during the sexua...

Figure 5.7. Structural organization of the LC ribozyme from Didymium iridis

Chapter 6

Figure 6.1. The different types of spontaneous reactions as a function of pH (...

Figure 6.2. Structure of the catalytic site of bovine ribonuclease A (pdb 6rsa...

Chapter 7

Figure 7.1. Phylogenetic tree showing the distribution by phylum and organelle...

Figure 7.2. Model of the events that may have led to the appearance of the LC ...

Figure 7.3. Potential mechanisms of hammerhead ribozymes based on their geneti...

Figure 7.4. Nucleolytic ribozymes are active as dimers

Figure 7.5. Hepatitis delta virus (HDV) ribozyme and a related human ribozyme ...

Figure 7.6. Ribozymes in the human genome

Guide

Cover Page

Table of Contents

Title Page

Copyright Page

Foreword

Preface

Begin Reading

References

Index

Other titles from ISTE in Biology and Biomedical Engineering

WILEY END USER LICENSE AGREEMENT

Pages

iii

iv

vii

viii

ix

x

xi

xii

xiii

1

2

3

4

5

6

7

8

9

10

11

12

13

14

15

16

17

18

19

20

21

22

23

24

25

26

27

28

29

30

31

32

33

34

35

36

37

38

39

40

41

42

43

44

45

46

47

48

49

50

51

52

53

54

55

56

57

58

59

60

61

62

63

64

65

66

67

68

69

70

71

72

73

74

75

76

77

78

79

80

81

82

83

84

85

86

87

88

89

90

91

92

93

94

95

96

97

98

99

100

101

102

103

104

105

106

107

108

109

110

111

112

113

114

115

116

117

118

119

120

121

122

123

124

125

126

127

128

129

130

131

132

133

134

135

136

137

138

139

140

141

142

143

144

145

146

147

148

149

150

151

152

153

154

155

156

157

158

159

160

161

162

163

164

165

166

167

168

169

170

171

168

Nucleic Acids Set

coordinated byMarie-Christine Maurel

Volume 2

Looking at Ribozymes

Biology of Catalytic RNA

Benoît Masquida

with the participation of Fabrice Leclerc

First published 2023 in Great Britain and the United States by ISTE Ltd and John Wiley & Sons, Inc.

Apart from any fair dealing for the purposes of research or private study, or criticism or review, as permitted under the Copyright, Designs and Patents Act 1988, this publication may only be reproduced, stored or transmitted, in any form or by any means, with the prior permission in writing of the publishers, or in the case of reprographic reproduction in accordance with the terms and licenses issued by the CLA. Enquiries concerning reproduction outside these terms should be sent to the publishers at the undermentioned address:

ISTE Ltd27-37 St George’s RoadLondon SW19 4EUUKwww.iste.co.uk

John Wiley & Sons, Inc.111 River StreetHoboken, NJ 07030USAwww.wiley.com

© ISTE Ltd 2023The rights of Benoît Masquida and Fabrice Leclerc to be identified as the authors of this work have been asserted by them in accordance with the Copyright, Designs and Patents Act 1988.

Any opinions, findings, and conclusions or recommendations expressed in this material are those of the author(s), contributor(s) or editor(s) and do not necessarily reflect the views of ISTE Group.

Library of Congress Control Number: 2023946400

British Library Cataloguing-in-Publication DataA CIP record for this book is available from the British LibraryISBN 978-1-78630-977-8

Foreword

Ribozymes are enzymes composed of RNA instead of proteins. Proteins have 20 different amino acid side chains, covering a wide range of aqueous chemistry. In contrast, RNA has fewer resources, with only four nucleic bases with a 2' hydroxyl group. However, its polyanionic nature allows it to recruit polycations, and thus bind hydrated metal ions that can also participate in chemical catalysis. Despite their limited catalytic resources, ribozymes are capable of accelerating a chemical reaction by a million times or more.

Until the 1980s, the only known biocatalysts were proteins. This all changed when Tom Cech discovered that splicing of the Tetrahymena group I intron was autocatalyzed. At the same time, Norm Pace and Sidney Altman were studying RNase P, which processes the 5' ends of RNA transfer, and they discovered that the active component was actually an RNA molecule, not a protein subunit. These discoveries opened up a whole new field of biological chemistry, challenging the chemist to understand how RNA could act as an enzyme with so few chemical resources.

Why are we interested in ribozymes? And why are they important in biology? First, they were most likely the only biological catalysts at an early stage in the development of life on Earth. There is a big “chicken and egg” problem in starting life, with both proteins and nucleic acids being needed. Evolution could not begin until there was genetic coding, replication and formation of biological catalysts. However, the RNA world hypothesis assumes that early life would have begun with RNA as both a genetic and catalytic reserve. This would therefore require ribozymes to catalyze primitive metabolism at this stage.

Proteins have a number of advantages over RNA as catalysts. It is therefore likely that the world of RNA would have been overtaken relatively quickly by a world of proteins. Nevertheless, ribozymes still exist in contemporary biology. This is the second reason why ribozymes are important. RNA catalyzes what is perhaps the most important reaction in the cell, the condensation of amino acids to form polypeptides in the large ribosomal subunit. That is, the ribosome is a ribozyme! The splicing of mRNA by the spliceosome is also catalyzed by RNA from the spliceosome. In addition, RNase P is involved in the maturation of tRNAs in all areas of life.

A group of nucleolytic ribozymes are the site-specific nucleases. These are generally part of the smaller catalytic RNA group. They were discovered in Australia, as they are present in viruses significant in the agricultural industry. In Adelaide, I met Bob Symons, who discovered the hammerhead ribozyme relatively late in life, and I thought he was more interested in grape growing than molecular biology! There are nine ribozymes in this class, the most recent of which were found through the application of directed bioinformatics in Ron Breaker’s lab. Some of the nucleolytic ribozymes have been found to be widespread, including in the human genome. In most cases, we do not yet know their function, but they are probably important.

This book presents a synthesis of the current state of knowledge in the field of ribozymes. So what are the big questions? Obviously, a major question is how can such major feats of chemical catalysis be achieved? This question can be asked at almost any level, and I am not sure we fully understand a protein enzyme. But in general terms, we have a general understanding, and for some ribozymes, our description of their catalytic mechanism is very good. Of course, any catalyst must ultimately stabilize the transition state of the reaction, but how do we achieve this? There are two major processes used by ribozymes, which divide them into two classes. Self-splicing introns and RNase P use hydrated metal ions to activate the nucleophile and organize the transition state. In contrast, nucleolytic ribozymes use general acid–base catalysis, with a major role for catalytic nucleobases.

A second major question is the extent to which RNA-based catalysis is used in current biology, and whether there are major classes of ribozymes that have escaped our discovery so far. Clearly, an RNA world would have required enzymes that catalyze a much wider range of chemical reactions, including “difficult” ones like C–C bond formation. It is very exciting to speculate on what solutions might exist, but how would we find them and where should we look? One way to find out what RNA might be capable of is to use in vitro selection methods. There has been a recent surge of interest in RNA species that can accelerate methyl transfer reactions, for example.

One way biology could have overcome the lack of chemical functionality of RNA might have been to use small molecules as cofactors, much like proteins use coenzymes. An example already exists with the nucleolytic ribozyme GlmS, which uses the captive ammonium group of glucosamine as the general acid. RNA is excellent at capturing small molecules, and here riboswitches can show the way. Many classes of riboswitches interact with potent coenzymes (suggesting an ancient origin tracing back to the RNA world), and we might imagine that such RNA molecules could relatively easily be converted into ribozymes using their coenzymes in catalysis.

Thus, ribozymes are important, and present a challenge to the chemist to understand their mechanisms, and at the same time perhaps shed a light on biocatalysis in general. In many cases, they also present interesting questions about their biology and the extent to which they have evolved in directions that we have not yet explored in depth. I hope that new generations will take up this challenge, and this book will provide an excellent starting point for that journey.

David M.J. LILLEY

University of Dundee

Preface

Ribozymes are enzymes composed of nucleic acids and in nature more specifically of RNA. They are at the origin of life as Thomas Cech and Sidney Altman proposed in an emblematic book entitled The RNA World: The Nature of Modern RNA Suggests a Prebiotic RNA World (Gesteland and Atkins 1993). The expansion of life indeed supposes the enzymatic copy of a pre-genetic material. Thus, nucleic acids combine two properties that proteins do not have. They pair via nucleobases by forming double helices in which one strand is a negative copy of the other, in reference to analog photography. In addition, the catalytic properties of their chemical groups, even if limited, accelerate millions of times over the transesterification reactions used for the synthesis of this copy. It is thus possible to design a self-replicating ribozyme that represents the archetype of an autonomous prebiotic system. The development of this primordial system led to the transmission and translation of the genetic material as we know it today, with the transcription of DNA into RNA followed by its translation into proteins.

In biology, there are numerous reminiscences of this system. The translation of messenger RNAs into proteins via the ribosome is probably the example that comes most immediately to mind since the ribosome is a ribozyme. Thus, the synthesis of the peptide bond of proteins, transpeptidylation, depends directly on RNA catalysis. The same is true for splicing by both autocatalytic introns, discovered by Thomas Cech, and spliced introns by the spliceosome. The vitamins and enzymatic cofactors of proteins are frequently nucleotide derivatives (FMN, FAD, NAD, vitamin B12, coenzyme A, ATP, SAM, to constitute a non-exhaustive list)1. These nucleotide derivatives specialized hundreds of millions of years ago upon contact with RNAs to perform specific chemical reactions beyond their reach, thus diversifying the catalytic repertoire of ribozymes and also leading to proteins emerging in the chemistry of life. This dependence of protein enzymes on these cofactors, many of which are derived from nucleic acids, as well as the relatively easy possibility of “selecting” in vitro RNA sequences capable of recognizing these same cofactors suggest that in the “RNA world” the catalytic diversity was greater than that visible today. This also means that the proteins offer reactional diversity, admittedly greater than that of RNAs, but still insufficient to do without these cofactors. Why would proteins have taken precedence over RNA at some point? Probably for reasons of genome stability and catalytic speed, but these are only partial answers. More recently identified ribozymes also suggest that they act in the developmental programs of organisms, cell differentiation, memory or stress. It is safe to say that our view of ribozymes in biology is sufficiently fragmentary that we have not yet grasped the diversity of their roles.

Hence, it became necessary to publish a book presenting this field of biology, in order to draw up an inventory allowing the greatest number of people to understand the importance of this group of molecules. Our ambition is to give the reader, from the beginning of higher education in life sciences to the confirmed researcher wishing to acquire structural knowledge, the possibility to quickly familiarize with this flourishing field that seems to have gone a bit out of fashion. However, this is not the case! Modern biology methods, and in particular high-throughput sequencing, are giving a second wind to ribozyme research. Indeed, since their discovery, the emblematic ribozymes have been mainly studied at a mechanistic level because their role was well established. But the presence of ribozymes in genomes points to many unsuspected roles that need to be studied further in order to gain a deeper understanding of the diversity and complexity of life. The preponderance of transcribed but untranslated sequences in genomes indicates an essential role for RNAs among which still unknown ribozymes certainly remain to be identified and studied.

This book covers four topics. Chapter 1 provides structural knowledge on nucleic acids in order to understand the following parts. Chapters 2 and 3describe the history of the discovery of ribozymes and review the state of knowledge about ribozymes, which have been studied since the 1980s. Catalysis by ribozymes is discussed in Chapters 4–6 and allows us to visualize the strategies used by these molecules to enable transesterification reactions. Finally, in Chapter 7, the roles of ribozymes are discussed in relation to the context in which they have been identified, thus whetting our curiosity when it is realized that more questions than answers are provided.

We hope you enjoy reading!

Acknowledgments

We thank Professor David M.J. Lilley (University of Dundee, United Kingdom) and Drs François Michel (Institut de systématique, évolution et biodiversité, Museum d’histoire naturelle, Paris) and Maria Costa (I2BC, Paris Saclay) for critically reviewing this manuscript. We would also like to thank Marie-Christine Maurel for giving us the opportunity to write this book and for her unfailing encouragement throughout the project.

More indirectly, we would also like to thank our mentors and researcher colleagues, as well as teacher-researchers and technicians who contribute to the advancement of knowledge. Research is a team effort where the notion of community is paramount. Community is understood here as a structure in which all members work in synergy with a common interest beyond particular or individual interests. Scientific research is a risk-taking activity in which even the instigators are unable to foresee the steps that will lead to discovery. This particular context can only foster discovery if researchers are free to focus their thoughts on their object, without these being polluted by materialistic considerations. “Research cannot be programmed, it takes time and cannot be done with an uncertain status,” to quote Rose Katz, a researcher at INSERM who passed away in 2022.

The three-dimensional molecule figures present in most of the images in this book were created using PyMol software (Schrodinger 2010).

September 2023

Note

1

FMN: flavin mononucleotide; FAD: flavin adenine dinucleotide; NAD: nicotinamide; ATP: adenosine triphosphate; SAM: S-adenosyl methionine.

1Fundamentals of RNA and Ribozyme Structure

This chapter is intended to familiarize the reader with the structure of RNAs. Understanding the structural basis of RNAs is a prerequisite for the study of ribozymes and RNA-mediated catalysis. This section shows how nucleotide stereochemistry guides the structuring of helices and consequently the addition of functional motifs that give this polymer its folding and interaction properties, as well as its catalytic properties.

It is important to understand that all biological mechanisms rely on the interaction capabilities of structured molecules. The structure of biomolecules is therefore a fundamental aspect for the understanding of biology. The figures in this book are therefore often developed from experimental structures obtained by radio-crystallography or electron microscopy. Visualizing a biological mechanism through the molecular structures involved allows a better understanding of the actions of the different partners and the domains that compose them. It is then possible to deduce the mechanisms of chemical reactions and also to establish evolutionary relationships between homologous molecules of different organisms that perpetuate these mechanisms while adapting to different selection pressures resulting from distinct ecological constraints.

1.1. Sequences and secondary structures

RNA (ribonucleic acid) is one of the three main biological polymers with DNA (deoxyribonucleic acid) and proteins. RNA adopts complex structures thanks to the physicochemical properties of the four main nucleotides from which it is assembled. The nucleotides are composed of a ribose-phosphate part and an aromatic part, the nucleobase, attached to the ribose. The base is composed of one or two fused aromatic rings containing imines and ethylenic carbons decorated by exocyclic amines and/or carbonyl groups. The ribose is also substituted with a phosphate group (Figure 1.1(a)). The phosphate group gives each nucleotide a negative charge. The “backbone” of the polymer is thus a polyanion. The decorations of the aromatic bases generate an electrostatic profile specific to each one that allows for the local appearance of negative (δ-) and/or positive (δ+) partial charges. RNAs are therefore not simple polyanions. The electrostatic profile of the bases confers on nucleotides interaction properties between them, as well as with the ions and water molecules that solvate them (Auffinger et al. 2016; D’Ascenzo and Auffinger 2016; Leonarski et al. 2017, 2019). Nucleotides tend to stack and form right-handed double-stranded helices promoted by the establishment of hydrogen bonds between bases. The 5'–3' orientation of the strands is opposite. The strands are therefore antiparallel (Figure 1.1(b)).

Figure 1.1.The four nucleotides of RNA form antiparallel double-stranded helical structures

COMMENTARY ON FIGURE 1.1.– a) Two purines (N) on the left and two pyrimidines on the right (Y). The ribose and the negatively charged phosphate group are shown on the adenine along with the numbering of the atoms according to the IUPAC (International Union of Pure and Applied Chemistry) system. The ribose atoms are numbered from 1' to 5' and the base atoms from 1 to 9 for a purine (R) and from 1 to 6 for a pyrimidine (Y). The oxygen atom of the ribose ring corresponds to the hydroxyl group carried by the C4' and therefore has the same number (O4'). The situation is identical for the hydroxyl groups carried by C2' and C3'. Uracil and cytosine have an O4 or N4 group, respectively. The polynucleic acid parent chains are oriented from 5' to 3'. The present example gives the sequence 5'-ABC-3'. b) Since each phosphate group carries a negative charge, a polynucleotide is a polyanion whose structure mostly forms right-handed antiparallel double-stranded helices. The bases pair up to form plateaus of bases that stack with each other.

Despite different electrostatic properties, the stereochemistry of each nucleotide is identical. The βD-ribofuranose isomer1, hereafter simply referred to as ribose, adopts an envelope (E) or twist (T) fold depending on whether four or three ring atoms define a plane, respectively. This plane is oriented using the only non-asymmetric carbon atom outside the ribose ring, the C5'. When the ribose is drawn with the C5' above the plane and to the left, the C2' and C3' carbon atoms point forward and the O4' points backward (Figure 1.2(a)). This configuration is the only one found in nature and gives an idea of the intensity of selection pressures that led to the emergence of these stereoisomers in biological nucleic acid synthesis pathways. The base is branched at the C1' position and points to the same edge as the phosphate group. All nucleotides are therefore superimposable to each other by their ribose-phosphate part. The strands (chains) of nucleotides are naturally structured into helices that interact with each other to form double-stranded helices characterized by grooves of different morphologies (Figure 1.2(b)). In fact, the path between the riboses of a base pair is shorter on one edge of the helix than on the other, which gives rise to the notion of major and minor grooves. In RNA, the minor groove is wide and shallow, and the major groove is narrow and deep. Width and depth are not independent. A deep groove is narrow, and a wide groove is shallow. The proteins that interact with RNA therefore tend to interact with the shallow groove.

Figure 1.2.Nucleotides and base pairs generating antiparallel double-stranded helices

COMMENTARY ON FIGURE 1.2.– a) Nucleotides adopt a precise conformation within the helices. The three edges of the nucleobases define the Watson–Crick, Hoogsteen and Sugar edges schematized by the edges of a right triangle. The atoms are numbered as shown. O1P and O2P are not linearly integrated into the 5'–3' ribose-phosphate backbone. The phosphate moiety is prochiral because the tetrahedral phosphorus has two identical moieties (O1P corresponds to the Pro-S stereoisomer and O2P corresponds to the Pro-R with reference to the R and S stereodescriptors). b) RNA helices are formed by the stacking of base pairs formed via Watson–Crick edges. Watson–Crick helices have an antiparallel orientation of the 5'–3' strands. If the 5' end of one strand is behind the plane of the sheet, then the 5' end of the other strand is above the plane. c) The permutations of these base pairs are isosteric, i.e. their riboses are superimposable 2 to 2. The orange dashed line collinear to the hydrogen bonds verifies whether the pairing is in the cis (both riboses are on the same edge) or trans configuration (both riboses are on opposite sides of this line). The O3'–P bonds between two adjacent riboses induce a right-handed rotation of about 33° between two consecutive base pairs. The A-form RNA helices therefore rotate to the right.

Five important properties of RNA follow from this absolute configuration.

The planar structure of nucleobases defines three hydrogen bond acceptor and/or donor edges, Watson–Crick (W or WC), Hoogsteen (H) and sugar (S). The W edge is responsible for the base pairs identified in the DNA double helix model by James Watson and Francis Crick in 1953 (Watson and Crick 1953). Karst Hoogsteen (1963) was the first to observe pairings involving the N7 and N6 positions of adenines, thus giving his name to this edge of the nucleobases:

– The O2' group plays a prominent role in interactions with the sugar edge and, as we will see later, in catalysis. Since DNA is free of the O2' group, its structural repertoire is consequently not as rich as that of RNA, as is its chemical reactivity.

– The nucleotides are linked to each other from the 5' position to the 3' position. The ribose-phosphate backbone is thus polarized. In projection on the axis of the helix, the nucleotide i-1 which precedes the nucleotide considered i is thus “above” the plane of the ribose.

– The plane of the base is perpendicular to the plane of the ribose and to the helix axis (Figure 1.2(b)).

– The grooves of the helix have different widths and depths. The deep groove is less accessible because it is narrow due to the stacking of the base plateaus. On the contrary, the shallow groove is very accessible. The S sides of the bases are therefore more accessible for another macromolecule than the H sides. The W sides are involved in pairing and are therefore less frequently involved in interactions with other molecular partners.

These five properties result in the formation of anti-parallel double-stranded helices. In a multi-nucleotide chain, the bases tend to stack with each other (stacking interactions), providing the edges capable of forming hydrogen bonds with the opportunity to interact with another RNA chain. For a helix to form, the interacting bases must be complementary. The Watson–Crick edges of a purine A or G always form a base pair with the Watson–Crick edges of a pyrimidine U or C, respectively, by establishing hydrogen bonds. In RNAs, G can also interact with U by forming two hydrogen bonds. This particular conformation is called wobble because it induces a shift towards the deep groove of the pyrimidine. This geometry stabilizes helices whose strands are not totally complementary. This geometry was proposed long before the first crystallographic structures by Crick (1966). This particular geometry is of great importance in biology both structurally and in terms of the genetic code that converts nucleic acid sequences into proteins, as we will see later.

In Watson–Crick base pairs, the riboses are on the same edge cis to a line collinear to the hydrogen bond axis (Figure 1.2(c)). This configuration results in the 5'–3' polarities of the strands of a helix being opposite. The helix is said to be antiparallel, and the positions of the elements of the ribose-phosphate backbone are always equivalent whatever the pairings A–U, U–A, G=C, C=G put in place. These pairings are said to be isosteric, i.e. of the same volume. Each pair of bases is therefore superimposable on the previous and the next, which allows them to be stacked. In addition, the βD stereoisomer of the ribose induces a right-hand twist of the helix of about 33° per helix turn (Figure 1.2(c)). Thus, the common part of the nucleotides forms the ribose-phosphate backbone at the periphery of the helix and the distinctive part that is the nucleobase forms the core. The nucleotides thus interact between them by their distinctive part, the nucleobase. The isostery between cisW