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Coordination Chemistry within the Core of Large RNAs: Regulating Tertiary Contacts and Function

Applicant Sigel Roland K. O.
Number 124834
Funding scheme Project funding (Div. I-III)
Research institution Institut für Chemie Universität Zürich
Institution of higher education University of Zurich - ZH
Main discipline Inorganic Chemistry
Start/End 01.04.2009 - 30.09.2012
Approved amount 483'860.00
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Keywords (7)

Metal Ions; RNA; NMR; ribozymes & riboswitches; ribozymes; bioinorganic chemistry

Lay Summary (English)

Lead
Lay summary
Metal ions are necessary for folding and function of catalytic RNA molecules. However, the structural and mechanistic roles of these ions are poorly understood. For living systems, it is usually assumed that Na(I) and Mg(II) ions are the sole metallic cofactors involved with nucleic acids, both being most abundant and freely available in the cell. However, in living organisms, a multitude of other metal ions is present, although tightly regulated. Furthermore, in biochemical experiments many different metal ions are applied. The recognition of specific metal ions by nucleic acids is very poorly understood as is their effect on structure, folding, and catalytic activity. This study focuses on the thermodynamic and structural characterization of these interactions in large RNAs, i.e. group II intron ribozymes and Mg(II) riboswitches. We are applying a multidisciplinary approach by using a combination of tools from Coordination and Analytical Chemistry as well as Structural Biology. Thus, our results will not only contribute to the understanding of the global structure and function of these naturally occurring RNAs, but they also promise to have a significant impact on the Bioinorganic Chemistry of RNAs and on RNA Biochemistry in general.
Direct link to Lay Summary Last update: 21.02.2013

Responsible applicant and co-applicants

Employees

Publications

Publication
NMR Spectroscopy in Bioinorganic Chemistry
Donghi D, Johannsen S, Sigel RKO, Freisinger E (2012), NMR Spectroscopy in Bioinorganic Chemistry, in CHIMIA, 66(10 - issue), 791-797.
Synthesis and Acid?Base Properties of an Imidazole-Containing Nucleotide Analog, 1-(2'-Deoxy-ss-D-ribofuranosyl)imidazole 5'-Monophosphate (dImMP2-)
Megger N, Johannsen S, Muller J, Sigel RKO (2012), Synthesis and Acid?Base Properties of an Imidazole-Containing Nucleotide Analog, 1-(2'-Deoxy-ss-D-ribofuranosyl)imidazole 5'-Monophosphate (dImMP2-), in CHEMISTRY & BIODIVERSITY, 9(9), 2050-2063.
Unusually High-Affinity Mg2+Binding at the AU-Rich Sequence within the Antiterminator Hairpin of a Mg2+Riboswitch
Korth MMT, Sigel RKO (2012), Unusually High-Affinity Mg2+Binding at the AU-Rich Sequence within the Antiterminator Hairpin of a Mg2+Riboswitch, in CHEMISTRY & BIODIVERSITY, 9(9), 2035-2049.
Accurate analysis of Mg2+ binding to RNA: From classical methods to a novel iterative calculation procedure
Erat MC, Coles J, Finazzo C, Knobloch B, Sigel RKO (2012), Accurate analysis of Mg2+ binding to RNA: From classical methods to a novel iterative calculation procedure, in COORDINATION CHEMISTRY REVIEWS, 256(1-2), 279-288.
MINAS-a database of Metal Ions in Nucleic AcidS
Schnabl J, Suter P, Sigel RKO (2012), MINAS-a database of Metal Ions in Nucleic AcidS, in NUCLEIC ACIDS RESEARCH, 40(D1), 434-438.
Characterization of Metal Ion-Nucleic Acid Interactions in Solution
Pechlaner Maria, Sigel Roland K. O. (2012), Characterization of Metal Ion-Nucleic Acid Interactions in Solution, in Metal Ions in Life Sciences, 10, 1-42.
Interplay between Metal Ions and Nucleic Acids
Sigel Astrid (Ed.), Sigel Helmut (Ed.), Sigel Roland K. O. (Ed.) (2012), Interplay between Metal Ions and Nucleic Acids, Springer, Dordrecht NL.
Metal Ion-RNA Interactions Studied via Multinuclear NMR
Donghi Daniela, Sigel Roland K. O. (2012), Metal Ion-RNA Interactions Studied via Multinuclear NMR, in Methods in Molecular Biology, 848, 253-274.
Probing the Metal-Ion-Binding Strength of the Hydroxyl Group
Al-Sogair FM, Operschall BP, Sigel A, Sigel H, Schnabl J, Sigel RKO (2011), Probing the Metal-Ion-Binding Strength of the Hydroxyl Group, in CHEMICAL REVIEWS, 111(8), 4964-5003.
Stability and Structure of Mixed-Ligand Metal Ion Complexes That Contain Ni2+, Cu2+, or Zn2+, and Histamine, as well as Adenosine 5 '-Triphosphate (ATP(4-)) or Uridine 5 '-Triphosphate (UTP4-): An Intricate Network of Equilibria
Knobloch B, Mucha A, Operschall BP, Sigel H, Jezowska-Bojczuk M, Kozlowski H, Sigel RKO (2011), Stability and Structure of Mixed-Ligand Metal Ion Complexes That Contain Ni2+, Cu2+, or Zn2+, and Histamine, as well as Adenosine 5 '-Triphosphate (ATP(4-)) or Uridine 5 '-Triphosphate (UTP4-): An Intricate Network of Equilibria, in CHEMISTRY-A EUROPEAN JOURNAL, 17(19), 5393-5403.
Structural and Catalytic Roles of Metal Ions in RNA
Sigel Astrid (Ed.), Sigel Helmut (Ed.), Sigel Roland K. O. (Ed.) (2011), Structural and Catalytic Roles of Metal Ions in RNA, RSC Publishing, Cambridge UK.
Metal Ions in Toxicology: Effects, Interactions, Interdependencies
Sigel Astrid (Ed.), Sigel Helmut (Ed.), Sigel Roland K. O. (Ed.) (2011), Metal Ions in Toxicology: Effects, Interactions, Interdependencies, RSC Publishing, Cambridge UK.
Methods to Detect and Characterize Metal Ion Binding Sites in RNA
Erat Michele C., Sigel Roland K. O. (2011), Methods to Detect and Characterize Metal Ion Binding Sites in RNA, in Metal Ions in Life Sciences, 9(37), 100.
Shaping RNA Structures with Metal Ions and Metal Ion Complexes
Sigel RKO, Gallo S (2010), Shaping RNA Structures with Metal Ions and Metal Ion Complexes, in CHIMIA, 64(3 - Alfred), 126-131.
A Stability Concept for Metal Ion Coordination to Single-Stranded Nucleic Acids and Affinities of Individual Sites
Sigel RKO, Sigel H (2010), A Stability Concept for Metal Ion Coordination to Single-Stranded Nucleic Acids and Affinities of Individual Sites, in ACCOUNTS OF CHEMICAL RESEARCH, 43(7), 974-984.
Metal ion-N7 coordination in a ribozyme branch domain by NMR
Erat MC, Kovacs H, Sigel RKO (2010), Metal ion-N7 coordination in a ribozyme branch domain by NMR, in JOURNAL OF INORGANIC BIOCHEMISTRY, 104(5), 611-613.
Controlling ribozyme activity by metal ions
Schnabl J, Sigel RKO (2010), Controlling ribozyme activity by metal ions, in CURRENT OPINION IN CHEMICAL BIOLOGY, 14(2), 269-275.
Solution structure of a DNA double helix with consecutive metal-mediated base pairs
Johannsen S, Megger N, Bohme D, Sigel RKO, Muller J (2010), Solution structure of a DNA double helix with consecutive metal-mediated base pairs, in NATURE CHEMISTRY, 2(3), 229-234.
Organometallics in Environment and Toxicology
Sigel Astrid (Ed.), Sigel Helmut (Ed.), Sigel Roland K. O. (Ed.) (2010), Organometallics in Environment and Toxicology, RSC Publishing, Cambridge UK.
Exploring Metal Ion Coordination to Nucleic Acids by NMR
Johannsen S, Korth MMT, Schnabl J, Sigel RKO (2009), Exploring Metal Ion Coordination to Nucleic Acids by NMR, in CHIMIA, 63(3 - Swiss ), 146-152.
The structural stabilization of the kappa three-way junction by Mg(II) represents the first step in the folding of a group II intron
Donghi Daniela, Pechlaner Maria, Finazzo Cinzia, Knobloch Bernd, Sigel Roland K. O., The structural stabilization of the kappa three-way junction by Mg(II) represents the first step in the folding of a group II intron, in Nucleic Acids Research, 00.

Collaboration

Group / person Country
Types of collaboration
Prof. Dr. Jens Müller, University of Münster Germany (Europe)
- in-depth/constructive exchanges on approaches, methods or results
- Publication
- Exchange of personnel
Dr. Helena Kovacs, Bruker Biospin, Fällanden (ZH) Switzerland (Europe)
- in-depth/constructive exchanges on approaches, methods or results
- Publication
- Research Infrastructure
Prof. Dr. Eva Freisinger, Universität Zürich Switzerland (Europe)
- in-depth/constructive exchanges on approaches, methods or results
- Research Infrastructure

Scientific events

Active participation

Title Type of contribution Title of article or contribution Date Place Persons involved
Fall Meeting of the Swiss Chemical Society 13.09.2012 ETH Zürich, Switzerland
11th European Biological Inorganic Chemistry Conference (EUROBIC11) 12.09.2012 Granada, Spain
40th International Conference on Coordination Chemistry, ICCC40 09.09.2012 Valencia, Spain
International Conference on Magnetic Resonance in Biological Systems (XXVth ICMRBS) 19.08.2012 Lyon, France
12th Eurasia Conference on Chemical Sciences (EuAsC2S-12), 16.04.2012 Corfu, Greece
Swiss RNA Workshop 03.02.2012 Bern, Switzerland
Zing Coordination Chemistry Conference 09.12.2011 Xcaret, Mexico
11th International Symposium on Applied Bioinorganic Chemistry (ISABC-11), 02.12.2011 Barcelona, Spain
XL National Congress on Magnetic Resonance, 26.09.2011 Parma, Italy
Fall Meeting of the Swiss Chemical Society, 09.09.2011 Lausanne, Switzerland
5th EuCheMS Conference on Nitrogen Ligands in Coordination Chemistry, Metalorganic Chemistry, Bioinorganic Chemistry, Materials and Catalysis, 04.09.2011 Granada, Spain
15th International Conference on Biological Inorganic Chemistry (ICBIC-15) 07.08.2011 Vancouver, Canada
Swiss RNA Workshop 2011 21.01.2011 University of Bern, Bern, Switzerland
Zing Nucleic Acids Conference 2010, 02.11.2010 Puerto Morelos, Mexico
Fall Meeting of the Swiss Chemical Society, 16.09.2010 Lausanne, Switzerland
XI International Symposium on Inorganic Biochemistry, 04.09.2010 Kudowa Zdrój, Poland
10th European Biological Inorganic Chemistry Conference (EuroBIC-10), 22.06.2010 Thessaloniki, Greece
10th International Symposium on Applied Bioinorganic Chemistry (ISABC 10) 25.09.2009 Debrecen, Hungary
COST D39 Conference: Metallo-Drug Design and Action 24.09.2009 Debrecen, Hungary
7th International NCCR Symposium on New Trends in Structural Biology 07.09.2009 Zürich, Switzerland
Fall Meeting of the Swiss Chemical Society 04.09.2009 Lausanne, Switzerland,
3rd European Conference on Chemistry for Life Sciences (3rd ECCLSc) 02.09.2009 Frankfurt, Germany
14th International Conference on Biological Inorganic Chemistry (ICBIC-14), 25.07.2009 Nagoya, Japan
Spanish Conference on Bioinorganic Chemistry (Reunión Cientifica de Bioinorgánica) 15.07.2009 Palma de Mallorac, Spain


Awards

Title Year
ERC Starting Grant 2010
Alfred Werner Preis 2009

Associated projects

Number Title Start Funding scheme
144964 Upgrade of the UZH NMR Core Facility 01.12.2012 R'EQUIP
192153 Principles of Metal-Ion-Assisted Folding and Structure of Functional RNAs 01.04.2020 Project funding (Div. I-III)
117999 Metal Ion-Sensing by Nucleic Acids in Nature and in the Test Tube 01.12.2007 Project funding (Div. I-III)
114759 Deciphering the Role of Metal Ions in RNAs 01.04.2007 SNSF Professorships
143750 Metal Ion-Guided Assembly and Structures of the Catalytic Core of Ribozymes 01.10.2012 Project funding (Div. I-III)

Abstract

SUMMARY AND CENTRAL AIMS OF THE STUDYMetal ions are necessary for folding and function of catalytic RNA molecules. However, the structural and mechanistic roles of these ions are poorly understood. For living systems, it is usually assumed that Na(I) and Mg(II) ions are the sole metallic cofactors involved with nucleic acids, both being most abundant and freely available in the cell. However, in living organisms, a multitude of other metal ions is present, although tightly regulated. Furthermore, in biochemical experiments many different metal ions are applied. The recognition of specific metal ions by nucleic acids is very poorly understood as is their effect on structure, folding, and catalytic activity. This study focuses on the thermodynamic and structural characterization of these interactions in large RNAs, i.e. group II intron ribozymes and Mg(II) riboswitches. We are applying a multidisciplinary approach by using a combination of tools from Coordination and Analytical Chemistry as well as Structural Biology. Thus, our results will not only contribute to the understanding of the global structure and function of these naturally occurring RNAs, but they also promise to have a significant impact on the Bioinorganic Chemistry of RNAs and on RNA Biochemistry in general.AIM A: Understanding the First Step of Group II Intron Folding Group II introns are among the largest occurring RNAs in Nature and are closely related to the eukaryotic spliceosomal machinery. These autocatalytic self-splicing introns exhibit a straight folding pathway to the active structure devoid of kinetic traps. Folding is initiated by Mg(II) coordination, whereby the kappa-zeta region within domain 1 (D1) is the key element for the first phase of folding. We will solve the NMR solution structure of this key region of the yeast mitochondrial Sc.ai5gamma intron and characterize the Mg(II) binding as well as the structural changes associated. Our results will lead to the detailed understanding of the first folding step, which guides the assembly of the remaining domains to the catalytically active structure. AIM B: Understanding the Main Step of Catalytic Core FormationThe kappa-zeta region exhibits a second crucial function in group II introns: It constitutes the main docking site for domain 5, comprising together the largest part of the catalytic core. We will determine the structure of the about 80 nucleotide long kappa-zeta/D5 complex by NMR, investigate the structural changes of the two (sub)domains, and characterize the crucial role of metal ions associated with this docking event. A special focus will be addressed to the different effects and binding modes of various metal ions, especially Mg(II) and Ca(II), as the latter ion strongly perturbs the folding pathway and thus inhibits splicing. Surface Plasmon Resonance spectroscopy (SPR) will allow us to characterize in detail the effect of these M(n+) on the D1-D5 docking, i.e. the final step of the catalytic core assembly.AIM C: Recognizing DNA for Retro-HomingThe 5'-splice site is recognized and defined by base pairing between the exon binding site 1 in domain 1 and the complementary intron binding site 1 on the 5'-exon. Alternatively, also a corresponding sequence on DNA can be recognized, thus initiating homing or transposition. A divalent metal ion has thereby been shown to promote the final structure of this recognition complex, presumably stabilizing a strong kink in the backbone just opposite the scissile phosphodiester. Based on a recently solved NMR structure of this d3'-EBS1·IBS1 complex, we will now determine the detailed coordination sphere of this crucial divalent metal ion and investigate its exact role in the splice site recognition as well as in the RNA·DNA complex standing at the beginning of the homing/retrotransposition pathway. Part of this work also includes the characterization of the influence of different metal ions on this binding event by SPR. Hence, our results will lead to the understanding of the role of metal ions in substrate recognition.AIM D: Metal Ion Coordination at Atomic ResolutionNucleic acids show a surprising selectivity and specificity for a given kind of metal ion. The binding of natural metal ions to nucleic acids is governed by fast ligand exchange as well as a combination of inner- and outer-sphere interactions, making the characterization of the metal ion-coordination sphere highly challenging. Aside from X-ray crystallography, we will use NMR to determine the binding pockets and the corresponding intrinsic metal ion-binding constants of the investigated RNAs as well as to elucidate the inner- and outer-sphere coordinating atoms. This will allow us to understand the coordination-chemical basis of the accelerating or inhibiting influence of metal ions in ribozymes as well as their structure-defining influence in all RNAs.AIM E: Triggering and Characterizing Structural Changes in a Mg(II) RiboswitchCytoplasmic Mg(II) levels are confined to a narrow range, whose regulation is poorly understood. It has recently been discovered that in bacteria like S. typhimurium or E. coli, two alternating stem-loop structures within a conserved riboswitch sequence in the 5'-UTR of the mgtA gene regulate the expression of a Mg(II) transporter by directly sensing Mg(II) in the cytoplasm. It is the goal of our study to investigate the alternating structures at low and high Mg(II) concentrations and their interconversion, to identify the Mg(II) binding sites, to determine the affinity constants as well as the intrinsic coordination chemistry responsible for the discrimination of Mg(II) from other divalent metal ions.To summarize, the results of this study will reveal metal ion-binding motifs and their direct roles in the function of two large RNAs. This characterization will provide an important basis for research related to either structure, folding, and/or catalysis of RNAs in general as well as for the biomedical application of group II introns and riboswitches.
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