Structure of human telomere G-quadruplex in the presence of a model drug along the thermal unfolding pathway

Abstract

A multi-technique approach, combining circular dichroism spectroscopy, ultraviolet resonance Raman spectroscopy and small angle scattering techniques, has been deployed to elucidate how the structural features of the human telomeric G-quadruplex d[A(GGGTTA)3GGG] (Tel22) change upon thermal unfolding. The system is studied both in the free form and when it is bound to Actinomycin D (ActD), an anticancer ligand with remarkable conformational flexibility. We find that at room temperature binding of Tel22 with ActD involves end-stacking upon the terminal G-tetrad. Structural evidence for drug-driven dimerization of a significant fraction of the G-quadruplexes is provided. When the temperature is raised, both free and bound Tel22 undergo melting through a multi-state process. We show that in the intermediate states of Tel22 the conformational equilibrium is shifted toward the (3+1) hybrid-type, while a parallel structure is promoted in the complex. The unfolded state of the free Tel22 is consistent with a self-avoiding random-coil conformation, whereas the high-temperature state of the complex is observed to assume a quite compact form. Such an unprecedented high-temperature arrangement is caused by the persistent interaction between Tel22 and ActD, which stabilizes compact conformations even in the presence of large thermal structural fluctuations.

Does Raman spectroscopy recognize different G‐quadruplex arrangements?

“…The decisive advantage of nonresonant Raman spectroscopy is its spectral richness and the ability to study nucleic acids samples in solution over a wide range of DNA concentrations, as a function of various physicochemical factors such as temperature or ionic strength, starting at concentrations typical of NMR measurements (and still accessible by CD spectroscopy) up to very high concentrations not readily accessible by other methods . Several publications recently also reported the use of resonant Raman spectroscopy employing ultraviolet excitation (Ultraviolet Resonance Raman), which, due to a strong enhancement of some vibrational modes (mainly corresponding to chromophoric bases), allows probing G‐quadruplexes at relatively low DNA concentrations (comparable with those found in ordinary CD and absorption measurements) but at the expense of lower spectral richness . Surface enhanced Raman scattering, which is based on orders of magnitude enhancement of Raman signal by strong electromagnetic field in the presence of appropriately designed metal nanoparticles, has been used to study ligand interaction or for detection of G‐quadruplexes .…”

Conformational stability of DNA in hydrated ionic liquid by synchrotron-based UV resonance raman

“…In the wavenumber region between 1300 to 1800 cm -1 , it is noteworthy the behaviour of the band centered at ~1487 cm -1 and labeled as band I. This peak is mainly attributable to guanine residues 29 arising from a combination of bending motion of C8-H and stretching movements of C8=N7 and N9-C8 (see Figure 1) 27,31 . increment of the intensity of the band I as a function of temperature both for DNA/IL = 1/22 (w/w) and for pristine DNA (see Inset of Figure 2).…”

“…4, it is possible to observe that the intensities of band I for the spectra of DNA in presence of IL are less intense with respect to those measured in pure DNA over the whole investigated temperature range. This hypochromic effect suggests that the addition of [C 4 MIM]Cl tends to favor the formation of a more compact structure in DNA doublestranded 29 . Moreover, the persistence of hypochromicity also after the unfolding of DNA gives indication that the base-stacking of guanines is quite effective for DNA in the presence of [C 4 MIM]Cl.…”

“…Such approach allows to disentangle in a very efficient way the contributions arising from individual bases in the spectra of DNA 28 . In particular, the choice of 250 nm as excitation wavelength gives the opportunity to enhance and isolate the ring inplane vibration associated to the guanine residues 29 . This Raman feature is highly sensitive to the thermal disordering and it can be used as molecular marker of thermal denaturation of DNA.…”

BMPQ-1 binds selectively to (3+1) hybrid topologies in human telomeric G-quadruplex multimers

“…The folding, formation, stabilization and dissolution of the human telomeric G-quadruplexes have been studied by a variety of structural, biophysical and chemical probe methods ( 14–19 ). Yet, there is limited data present on the structural arrangements of G-quadruplex long repeats.…”