Interactions of the Human Telomere Sequence with the Nanocavity of the α-Hemolysin Ion Channel Reveal Structure-Dependent Electrical Signatures for Hybrid Folds

An, Na; Fleming, Aaron M.; Burrows, Cynthia J.

doi:10.1021/ja400973m

Cited by 49 publications

(107 citation statements)

References 61 publications

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“…Previously, we identified four different conformations that the human telomere sequence adopts in an aqueous KCl solution: hybrid-1 ( Fig. 2A), hybrid-2, and their corresponding triplexes with the double-chain reversal loop unfolded (33), in agreement with other singlemolecule studies and computational work (34,35). Current-time (i-t) traces of the events that correlated with the hybrid-folded DNA showed intermediate current levels (I M = 37% or 44%; Fig.…”

Section: Resultssupporting

confidence: 86%

“…Consistent with our previous investigation, translocation events indicating the existence of the triplexes also were observed in ∼19% of the total events, and they could be compared with the control sequence, in which a 5′-GGG sequence was replaced with 5′-TTT to enforce the formation of the triplex (SI Appendix, Fig. S9) (33).…”

Section: Resultssupporting

confidence: 86%

“…When these overhangs were removed, only the 37% current blockage level was observed, consistent with the observation that the hybrid-1 structure predominates when the dangling ends are missing (7,36). At the end of each of the events, the current level was observed to be at an intermediate level I M immediately before returning to the open-channel current; we interpreted this to mean that hybrid-folded oligomers were too stable inside the vestibule to unravel and translocate, and the DNA exited the vestibule from the cis opening where it had entered (33). Overall, the previous study showed that the α-HL vestibule might serve as a sieve to trap stably folded hybrid-1 and hybrid-2 structures and to report on their dynamic population in bulk solution.…”

Section: Resultssupporting

confidence: 84%

“…S5), indicating the spontaneous nature of the unraveling process. The possibility that these two duration populations resulted from the presence of hybrid-1 vs. hybrid-twofold was tested by examining the translocation events of 8-BrGsubstituted sequences, which selectively induced either the hybrid-1 or hybrid-2 conformations, depending on the location of the purines locked into the syn conformation (33). Similar event durations were observed for both sequences, ruling out such a possibility.…”

Section: Resultsmentioning

confidence: 99%

“…Our previous work demonstrated the ability of α-HL to resolve two very similar but highly dynamic and interchangeable folds: hybrid-1 and hybrid-2 (33). Herein, we examined a comprehensive library of proposed nanostructures derived from the human telomere sequence to explore the potential challenge posed to sequencing efforts, and the capability of α-HL to study a larger range of nanostructures to a deeper extent.…”

Section: Significancementioning

confidence: 99%

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Single-molecule investigation of G-quadruplex folds of the human telomere sequence in a protein nanocavity

Fleming

Middleton

et al. 2014

Proc. Natl. Acad. Sci. U.S.A.

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101

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Human telomeric DNA consists of tandem repeats of the sequence 5′-TTAGGG-3′ that can fold into various G-quadruplexes, including the hybrid, basket, and propeller folds. In this report, we demonstrate use of the α-hemolysin ion channel to analyze these subtle topological changes at a nanometer scale by providing structuredependent electrical signatures through DNA-protein interactions. Whereas the dimensions of hybrid and basket folds allowed them to enter the protein vestibule, the propeller fold exceeds the size of the latch region, producing only brief collisions. After attaching a 25-mer poly-2′-deoxyadenosine extension to these structures, unraveling kinetics also were evaluated. Both the locations where the unfolding processes occur and the molecular shapes of the G-quadruplexes play important roles in determining their unfolding profiles. These results provide insights into the application of α-hemolysin as a molecular sieve to differentiate nanostructures as well as the potential technical hurdles DNA secondary structures may present to nanopore technology.α-hemolysin nanopore | single-molecule detection N ucleic acids can fold into a myriad of secondary structures that depend on the primary sequence as well as the physical conditions in which the structures are prepared and characterized. One prime example of a multistructural sequence is human telomeric DNA comprising the repeat sequence 5′-TTAGGG-3′. This guanine-rich single-stranded sequence is known to fold into highly ordered nanostructures in the form of G-quadruplexes that feature the coordination of two alkali cations to three layers of G-tetrads formed by Hoogsteen hydrogen-bonded assemblies of four guanine bases ( Fig. 1A) (1, 2). G-quadruplexes provide a fascinating case in which the cation and physical context play critical roles in defining the overall structural topology as well as the stability of the fold. Guanine-rich sequences also are known to present challenges to PCR amplification and sequencingby-synthesis methods that require processive analysis of singlestranded DNA (ssDNA) because of the high thermodynamic stability of these folded structures.The following studies highlight the cation and contextdependent conditions in which the topology of the natural human telomere sequence 5′-TAGGG(TTAGGG) 3 TT-3′ is affected. In NaCl solution, NMR studies revealed a structure referred to as the basket fold (Fig. 1A) (4). Key features of this structure include an antiparallel strand arrangement with alternating syn and anti orientations of the guanosine glycosidic bonds and three tetrads linked by two edgewise loops and one diagonal loop. In contrast, NMR-based studies in KCl solution show that the same sequence folds predominantly to the hybrid-1 and hybrid-2 structures (Fig. 1A) (5-8). Characteristics of this structure are an antiparallel strand orientation with a 3+1 core of syn and anti G nucleotides yielding three tetrads. The loop topology of the hybrid fold consists of a double-chain reversal loop and two edgewise loops, in which hybr...

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Section: Resultssupporting

confidence: 86%

Section: Resultssupporting

confidence: 86%

Section: Resultssupporting

confidence: 84%

Section: Resultsmentioning

confidence: 99%

Section: Significancementioning

confidence: 99%

See 3 more Smart Citations

Single-molecule investigation of G-quadruplex folds of the human telomere sequence in a protein nanocavity

Fleming

Middleton

et al. 2014

Proc. Natl. Acad. Sci. U.S.A.

Self Cite

101

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Direct and Single‐Molecule Visualization of the Solution‐State Structures of G‐Hairpin and G‐Triplex Intermediates

et al. 2014

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We present the direct and single-molecule visualization of the in-pathway intermediates of the G-quadruplex folding that have been inaccessible by any experimental method employed to date. Using DNA origami as a novel tool for the structural control and high-speed atomic force microscopy (HS-AFM) for direct visualization, we captured images of the unprecedented solution-state structures of a tetramolecular antiparallel and (3+1)-type G-quadruplex intermediates, such as G-hairpin and G-triplex, with nanometer precision. No such structural information was reported previously with any direct or indirect technique, solution or solid-state, single-molecule or bulk studies, and at any resolution. Based on our results, we proposed a folding mechanism of these G-quadruplexes.

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Identifying and Differentiating Topological G‐Quadruplex Structures with DNA‐Encoded Plasmonic Gold Nanoparticles

et al. 2022

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DNA G‐quadruplexes (G4s) have been identified as critical elements in modulating genomic functions and many other biological processes. Their functions are highly dependent on the primary nucleotides and secondary folding structures. Therefore, to understand their functions, methods to identify and differentiate structures of G4 with speed and accuracy are required but limited. In this report, we have applied a synthetic G4 DNA‐encoded nanoparticle approach to identify and differentiate G4 DNA molecules with different topologies and nucleotide residues. We found that the resulting plasmonic properties of the gold nanoparticles, monitored by UV/Vis spectroscopy, are quite sensitive to different G4 structures, including stacking layers, loop sequences, capping bases on G4s, and topological structures. Through these systematic investigations, we demonstrate that this G4‐encoded gold nanoparticle approach can be used to profile the G4 structures and distinguish G4s from human telomeres. Such a method may have wide applications in G4 research.

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Interactions of the Human Telomere Sequence with the Nanocavity of the α-Hemolysin Ion Channel Reveal Structure-Dependent Electrical Signatures for Hybrid Folds

Cited by 49 publications

References 61 publications

Single-molecule investigation of G-quadruplex folds of the human telomere sequence in a protein nanocavity

Single-molecule investigation of G-quadruplex folds of the human telomere sequence in a protein nanocavity

Direct and Single‐Molecule Visualization of the Solution‐State Structures of G‐Hairpin and G‐Triplex Intermediates

Identifying and Differentiating Topological G‐Quadruplex Structures with DNA‐Encoded Plasmonic Gold Nanoparticles

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