Characterization of G-Quadruplexes Folding/Unfolding Dynamics and Interactions with Proteins from Single-Molecule Force Spectroscopy

Abstract

G-quadruplexes (G4s) are stable secondary nucleic acid structures that play crucial roles in many fundamental biological processes. The folding/unfolding dynamics of G4 structures are associated with the replication and transcription regulation functions of G4s. However, many DNA G4 sequences can adopt a variety of topologies and have complex folding/unfolding dynamics. Determining the dynamics of G4s and their regulation by proteins remains challenging due to the coexistence of multiple structures in a heterogeneous sample. Here, in this mini-review, we introduce the application of single-molecule force–spectroscopy methods, such as magnetic tweezers, optical tweezers, and atomic force microscopy, to characterize the polymorphism and folding/unfolding dynamics of G4s. We also briefly introduce recent studies using single-molecule force spectroscopy to study the molecular mechanisms of G4-interacting proteins.

Role of fast dynamics in the complexation of G-quadruplexes with small molecules

“…From these values, we estimated the resulting unfolding forces F u by applying the Lindemann criterion (eqn (7) and (8)), as shown in Table 1. Quite interestingly, they are of the same order of magnitude of the unfolding forces determined from single-molecule pulling experiments, 53,54 thus suggesting that the biomolecule hypersurface potential energy determines both the equilibrium G4 fast dynamics and the out-of-equilibrium mechanical unfolding experiments.…”

“Dynamical Docking” of Cyclic Dinuclear Au(I) Bis-N-heterocyclic Complexes Facilitates Their Binding to G-Quadruplexes

“…The G4-stabilizing properties of the isolated bis-NHC Au­(I) complexes, AuB2 , AuB3 , and AuC1–3 , were assessed via the FRET DNA melting assay using a panel of five G4s in a 5:1 stoichiometry [Au­(I) NHC complex: G4, see the Experimental Section for details]. The G4s were chosen to represent both the telomeric (hTelo) and promoter ( cKIT1 , cKIT2 , BCL2 , and hTERT ) regions of the polynucleotide, as well as different G4 strand orientations such as hybrid-mixed (3 + 1) (hTelo and BCL2 ) and parallel ( cKIT1 , cKIT2 , and hTERT ). − The bis-caffeine complex AuTMX 2 was tested as a benchmark, also because its cKIT2 and BCL2 G4-stabilizing properties have never been reported so far. , …”

Spontaneous DNA Synapsis by Forming Noncanonical Intermolecular Structures

“…However, synaptic complex formation also reduces duplex stability which may contribute to synaptic complex folding. In vivo conditions (≈100 mM KCl, molecular crowding) stabilise duplexes and synaptic complexes [ 55 , 56 , 57 ]. Synaptic complex folding requires DNA duplex melting.…”