Positional Impact of Fluorescently Modified G-Tetrads within Polymorphic Human Telomeric G-Quadruplex Structures

Abstract

Emissive C8-aryl-2'-deoxyguanosines placed within G-tetrads of G-quadruplex structures are useful probes for distinguishing G-quadruplexes from duplex structures using fluorescence spectroscopy. Here, we report the positional impact of C8-furyl-dG ((Fur)dG) on G-quadruplex folding in the human telomere 22-mer oligonucleotide (HTelo22, (d[AG3(T2AG3)3])). The (Fur)dG probe was inserted into four different positions within the three unique G-tetrads of HTelo22, and G-quadruplex folding was monitored by UV-vis thermal denaturation, circular dichroism, and fluorescence spectroscopy. Our studies demonstrate the impacts of C8-aryl-dG adduct formation on G-quadruplex polymorphism in K(+) solution and in the presence of the additives and cosolutes, CH3CN, polyethylene glycol (PEG-600), and N-methyl mesoporphyrin IX (NMM). Our experiments predict that C8-aryl-dG derivatives can serve as useful tools for various in vitro studies aimed at understanding both the implications of DNA adduct formation within G-quadruplex structures and the unique implications imposed by various folding topologies on biological function/recognition.

Conformation-sensitive nucleoside analogues as topology-specific fluorescence turn-on probes for DNA and RNA G-quadruplexes

“…Phosphoramidite 3 was used in the synthesis of fluorescent h-Telo DNA ONs 5 and 6 in which the loop residues dT 11 and dT 12 , respectively, were replaced with 5-benzofuran-modified 2′-deoxyuridine 1 (Figure 2 , Supplementary Data). We purposely chose to modify the loop dT residues as modifications on guanosine could affect the efficiency of formation of GQ ( 67 , 78 ). The integrity of full-length modified h-Telo DNA ONs 5 and 6 was confirmed by mass and HPLC analyses (Supplementary Figure S1 and Supplementary Table S2).…”

“…Moreover, some of the easily accessible analogues (e.g. 2-AP, 6-MI, furyl-dG), though exhibit enhancement in fluorescence depending on the position of incorporation, significantly destabilize the GQ structures ( 66 , 67 , 78 ). Hence, implementation of such probes in screening formats to identify efficient GQ binders may not be feasible.…”

“…Alternatively, fluorescent purine surrogates (e.g. 6-methylisoxanthopterin and 2-aminopurine) and base-modified 2′-deoxyguanosine analogues containing vinyl, styryl, aryl or heteroaryl group have been incorporated into ONs and utilized in the study of DNA GQs ( 58 – 67 ). However, many of the fluorescent non-covalent binders and nucleoside analogues are structurally perturbing or poorly discriminate different GQ topologies or exhibit unfavourable photophysical properties (e.g.…”

Dual fluorescent deoxyguanosine mimics for FRET detection of G-quadruplex folding

“…15 However, in antiparallel GQ structures they can occupy syn-G positions without disturbing H-bonding interactions. 16 Based on our previous studies using fluorescent 8aryldG bases to monitor GQ formation, [17][18][19] the emissive properties of Fur dG (Fig. 1a) .…”

“…This observation can be explained by the reduced quantum yield of Fur dG (D) in the B-form duplex. [17][18][19] Given that the A emission intensity in the GQ could be clearly distinguished from its emission intensity in the duplex following D excitation at 315 nm (Fig. 2b), the ability of the mTBA sample to signal thrombin binding through FRET was determined.…”