Folding thermodynamics of the hybrid‐1 type intramolecular human telomeric G‐quadruplex

Abstract

Guanine-rich DNA sequences that may form G-quadruplexes are located in strategic DNA loci with the ability to regulate biological events. G-quadruplexes have been under intensive scrutiny owing to their potential to serve as novel drug targets in emerging anticancer strategies. Thermodynamic characterization of G-quadruplexes is an important and necessary step in developing predictive algorithms for evaluating the conformational preferences of G-rich sequences in the presence or the absence of their complementary C-rich strands. We use a combination of spectroscopic, calorimetric, and volumetric techniques to characterize the folding/unfolding transitions of the 26-meric human telomeric sequence d[A3G3(T2AG3)3A2]. In the presence of K+ ions, the latter adopts the hybrid-1 G-quadruplex conformation, a tightly packed structure with an unusually small number of solvent-exposed atomic groups. The K+-induced folding of the G-quadruplex at room temperature is a slow process that involves significant accumulation of an intermediate at the early stages of the transition. The G-quadruplex state of the oligomeric sequence is characterized by a larger volume and compressibility and a smaller expansibility than the coil state. These results are in qualitative agreement with each other all suggesting significant dehydration to accompany the G-quadruplex formation. Based on our volume data, 432±19 water molecules become released to the bulk upon the G-quadruplex formation. This large number is consistent with a picture in which DNA dehydration is not limited to water molecules in direct contact with the regions that become buried but involves a general decrease in solute-solvent interactions all over the surface of the folded structure.

Structure, properties, and biological relevance of the DNA and RNA G-quadruplexes: Overview 50 years after their discovery

“…In the view of authors, the G4 forma tion follows a very complex kinetic pathway involving multiple alternative states, and only a small fraction of molecules is folded into the final G4 structure. Data obtained recently by NMR [159] and CD [110] agree with such kinetic pattern.…”

“…THERMODYNAMICS AND KINETICS OF QUADRUPLEX FOLDING-UNFOLDING To characterize G quadruplexes formed in a certain genome locus, each identified G4 motif is investigated in vitro under different experimental conditions close, to a lesser or greater degree, to the intracellular ones. The methods commonly used for the description of G4 topol ogy and determination of thermodynamic and kinetic parameters of G quadruplex folding and unfolding include CD [106,107] and UV spectroscopy [108,109], differential scanning calorimetry [110,111], gel elec trophoresis [29,52], X ray crystallography [25,112], chemical and enzymatic probing [101,113,114], fluores cence spectroscopy, fluorescence resonance energy trans fer (FRET) [34,46,115,116], including single molecule BIOCHEMISTRY (Moscow) Vol. 81 No.…”

“…The thermodynam ic contribution of dehydration and capture of counter ions was estimated in [50]. It was shown that water affect ed thermodynamic stability and polymorphism of G4s [110,144]. The decrease in water activity caused by addi tion of various alcohols or polyethylene glycol (PEG) into aqueous solution, results in stabilization of the quadru plex [145]; PEG is usually used as molecular crowding agent to create cell mimicking conditions.…”

The Deep Sea Osmolyte Trimethylamine N‐Oxide and Macromolecular Crowders Rescue the Antiparallel Conformation of the Human Telomeric G‐Quadruplex from Urea and Pressure Stress

“…In this study, we set out to explore the combined effects of HHP and common co‐solvents such as trimethylamine N ‐oxide (TMAO), urea, and the macromolecular crowding agent Ficoll on the conformational landscape of the human telomeric oligonucleotide sequence [A(GGGTTA) 3 GGG]. The conformations of G4‐DNAs are generally investigated using ensemble techniques such as circular dichroism (CD), UV/Vis, and NMR spectroscopy . The single‐molecule fluorescence resonance energy transfer (smFRET) technique as used herein allows to distinguish different folded conformations .…”

“…[16] In this study,w es et out to explore the combined effects of HHP and commonc o-solvents such as trimethylamine N-oxide (TMAO), urea, and the macromolecular crowding agent Ficoll on the conformational landscapeo ft he human telomeric oligonucleotide sequence [A(GGGTTA) 3 GGG].T he conformations of G4-DNAs are generally investigated using ensemble techniques such as circulard ichroism (CD), UV/Vis, and NMR spectroscopy. [4,5,[17][18][19][20][21][22] The single-molecule fluorescence resonance energy transfer (smFRET) technique as used herein allows to distinguish different folded conformations. [23][24][25][26] Recently,w ew ere also able to setup aH HP smFRET stage ( Figure S1 in the Supporting Information) allowing us to reach pressures up tõ 2kbar (200 MPa), covering the whole relevant pressure range on Earth.…”