Nejvíce citovaný článek - PubMed ID 15339186
Guanine quadruplex (GQ) is a noncanonical nucleic acid structure formed by guanine-rich DNA and RNA sequences. Folding of GQs is a complex process, where several aspects remain elusive, despite being important for understanding structure formation and biological functions of GQs. Pulling experiments are a common tool for acquiring insights into the folding landscape of GQs. Herein, we applied a computational pulling strategy─steered molecular dynamics (SMD) simulations─in combination with standard molecular dynamics (MD) simulations to explore the unfolding landscapes of tetrameric parallel GQs. We identified anisotropic properties of elastic conformational changes, unfolding transitions, and GQ mechanical stabilities. Using a special set of structural parameters, we found that the vertical component of pulling force (perpendicular to the average G-quartet plane) plays a significant role in disrupting GQ structures and weakening their mechanical stabilities. We demonstrated that the magnitude of the vertical force component depends on the pulling anchor positions and the number of G-quartets. Typical unfolding transitions for tetrameric parallel GQs involve base unzipping, opening of the G-stem, strand slippage, and rotation to cross-like structures. The unzipping was detected as the first and dominant unfolding event, and it usually started at the 3'-end. Furthermore, results from both SMD and standard MD simulations indicate that partial spiral conformations serve as a transient ensemble during the (un)folding of GQs.
- MeSH
- biomechanika MeSH
- DNA chemie MeSH
- G-kvadruplexy * MeSH
- mechanické jevy MeSH
- simulace molekulární dynamiky * MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Názvy látek
- DNA MeSH
Recently, we reported an inhibitory effect of guanine substitutions on the conformational switch from antiparallel to parallel quadruplexes (G4) induced by dehydrating agents. As a possible cause, we proposed a difference in the sensitivity of parallel and antiparallel quadruplexes to the guanine substitutions in the resulting thermodynamic stability. Reports on the influence of guanine substitutions on the biophysical properties of intramolecular parallel quadruplexes are rare. Moreover, such reports are often complicated by the multimerisation tendencies of parallel quadruplexes. To address this incomplete knowledge, we employed circular dichroism spectroscopy (CD), both as stopped-flow-assisted fast kinetics measurements and end-point measurements, accompanied by thermodynamic analyses, based on UV absorption melting profiles, and electrophoretic methods. We showed that parallel quadruplexes are significantly more sensitive towards guanine substitutions than antiparallel ones. Furthermore, guanine-substituted variants, which in principle might correspond to native genomic sequences, distinctly differ in their biophysical properties, indicating that the four guanines in each tetrad of parallel quadruplexes are not equal. In addition, we were able to distinguish by CD an intramolecular G4 from intermolecular ones resulting from multimerisation mediated by terminal tetrad association, but not from intermolecular G4s formed due to inter-strand Hoogsteen hydrogen bond formation. In conclusion, our study indicates significant variability in parallel quadruplex structures, otherwise disregarded without detailed experimental analysis.
- Klíčová slova
- DNA secondary structure, circular dichroism, multimerisation, parallel guanine quadruplex, stopped-flow,
- MeSH
- cirkulární dichroismus MeSH
- DNA chemie genetika MeSH
- G-kvadruplexy MeSH
- guanin chemie MeSH
- konformace nukleové kyseliny MeSH
- molekulární modely MeSH
- substituce aminokyselin * MeSH
- termodynamika MeSH
- vodíková vazba MeSH
- Publikační typ
- časopisecké články MeSH
- Názvy látek
- DNA MeSH
- guanin MeSH
G-quadruplexes are noncanonical nucleic acid structures formed from stacked guanine tetrads. They are frequently used as building blocks and functional elements in fields such as synthetic biology and also thought to play widespread biological roles. G-quadruplexes are often studied as monomers, but can also form a variety of higher-order structures. This increases the structural and functional diversity of G-quadruplexes, and recent evidence suggests that it could also be biologically important. In this review, we describe the types of multimeric topologies adopted by G-quadruplexes and highlight what is known about their sequence requirements. We also summarize the limited information available about potential biological roles of multimeric G-quadruplexes and suggest new approaches that could facilitate future studies of these structures.
- Klíčová slova
- DNA:RNA hybrid, G-quadruplex, R-loop, dimer, multimer, oligomer, promoter, telomere, tetramer,
- MeSH
- DNA chemie MeSH
- G-kvadruplexy * MeSH
- konformace nukleové kyseliny * MeSH
- molekulární modely MeSH
- molekulární struktura MeSH
- RNA chemie MeSH
- telomery MeSH
- vztahy mezi strukturou a aktivitou MeSH
- Publikační typ
- časopisecké články MeSH
- přehledy MeSH
- Názvy látek
- DNA MeSH
- RNA MeSH
G-quadruplexes can multimerize under certain conditions, but the sequence requirements of such structures are not well understood. In this study, we investigated the ability of all possible variants of the central tetrad in a monomeric, parallel-strand G-quadruplex to form higher-order structures. Although most of these 256 variants existed primarily as monomers under the conditions of our screen, ∼10% formed dimers or tetramers. These structures could form in a wide range of monovalent and divalent metal ions, and folding was highly cooperative in both KCl and MgCl2. As was previously shown for G-quadruplexes that bind GTP and promote peroxidase reactions, G-quadruplexes that form dimers and tetramers have distinct sequence requirements. Some mutants could also form heteromultimers, and a second screen was performed to characterize the sequence requirements of these structures. Taken together, these experiments provide new insights into the sequence requirements and structures of both homomultimeric and heteromultimeric G-quadruplexes.
