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Exploring the Formation Pathways of DNA G-Quadruplex Architectures
Rosu, Frédéric; Poncelet, Harmonie; Teulade-Fichou, Marie-Paule et al.
20072nd European Conference on Chemistry for Life Sciences
 

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Keywords :
mass spectrometry; G-quadruplex; electrospray; noncovalent; affinity; stoichiometry; binding
Abstract :
[en] Guanine-rich DNA strands can form the so-called G-quadruplex architectures due to the formation of quartets of guanines linked by 8 hydrogen bonds. G-quadruplexes are further stabilized by the inclusion of cations between the G-quartets. The abundance of G-rich regions throughout the genome and their very presence in telomeric regions made G-quadruplexes interesting targets. NMR and crystallographic studies of G-quadruplex structures revealed amazing variety in the G-quadruplex topologies. The next challenge will be to understand the rules governing the formation of the various topologies, in order to predict relevant G-quadruplexes in the genome, and in order to act rationally on their formation or disruption. To date, only few experimental [1] or theoretical [2] studies have been devoted to investigating the mechanisms of G-quadruplex formation. We report here a detailed investigation of DNA G-quadruplex formation pathways using electrospray mass spectrometry (ESI-MS). The sequences TGnT (n = 3-6) were purchased from Eurogentec (Seraing, Beliugm). ESI-MS experiments were performed in the negative ion mode on a Q-TOF Ultima Global (Waters, Manchester, UK). The cation used was ammonium (up to 150 mM). Experiments were performed in the presence and absence of methanol (up to 20%) as co-solvent. ESI-MS allows counting both the number of strands and the number of cations in each intermediate. We could confirm the presence of transient dimer and trimer intermediates in low abundance. More unexpectedly, ESI-MS also reveals unambiguously the formation of pentamers which contain ammonium cations. The pentamers slowly convert into tetramers. Counting the number of included cations also revealed that, in the case of (TG6T)4, inclusion of four ammonium cations is fast, while the inclusion of the last ammonium ion is very slow. We also found that the addition of methanol (initially added to obtain higher ion intensities) significantly increases the rate of G-quadruplex formation. Finally, we also investigated the role of G-quadruplex ligands in the rate of formation of G-quadruplexes. We could classify the ligands according to their increase of G-quadruplex formation kinetics, and distinguish the intermediates. Interestingly, one ligand showed formation of a higher-order structure by bridging two G-quadruplexes. Acknowledgement: The authors thank the FRS-FNRS for their support. References: [1] J. Gros et al., Nucleic Acids Res., 2007, doi:10.1093/nar/gkm111. [2] R. Stefl et al., Biophys. J., 2003, 85(3), 1787-1804.
Research center :
Giga-Systems Biology and Chemical Biology - ULiège
Disciplines :
Biochemistry, biophysics & molecular biology
Chemistry
Author, co-author :
Rosu, Frédéric ;  Université de Liège - ULiège > Département de chimie (sciences) > GIGA-R : Laboratoire de spectrométrie de masse (L.S.M.)
Poncelet, Harmonie;  Université de Liège - ULiège > Département de Chimie (Sciences) > GIGA-R : Laboratoire de Spectrométrie de Masse
Teulade-Fichou, Marie-Paule;  Collège de France, Paris
De Pauw, Edwin  ;  Université de Liège - ULiège > Département de chimie (sciences) > GIGA-R : Laboratoire de spectrométrie de masse (L.S.M.)
Gabelica, Valérie ;  Université de Liège - ULiège > Département de chimie (sciences) > GIGA-R : Laboratoire de spectrométrie de masse (L.S.M.)
Language :
English
Title :
Exploring the Formation Pathways of DNA G-Quadruplex Architectures
Publication date :
07 September 2007
Event name :
2nd European Conference on Chemistry for Life Sciences
Event place :
Wroclaw, Poland
Event date :
September 4-7, 2007
By request :
Yes
Audience :
International
Funders :
F.R.S.-FNRS - Fonds de la Recherche Scientifique [BE]
Available on ORBi :
since 11 February 2011

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