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G-rich VEGF aptamer with locked and unlocked nucleic acid modifications exhibits a unique G-quadruplex fold.

Marušič M, Veedu RN, Wengel J, Plavec J - Nucleic Acids Res. (2013)

Bottom Line: Both 5' with 3 nt and 3' with 4 nt overhangs display well-defined conformations, with latter adopting a basket handle topology.Locked residues contribute to thermal stabilization of the adopted structure and formation of structurally pre-organized intermediates that facilitate folding into a single G-quadruplex.Understanding the impact of chemical modifications on folding, thermal stability and structural polymorphism of G-quadruplexes provides means for the improvement of vascular endothelial growth factor aptamers and advances our insights into driving nucleic acid structure by locking or unlocking the conformation of sugar moieties of nucleotides in general.

View Article: PubMed Central - PubMed

Affiliation: Slovenian NMR Center, National Institute of Chemistry, Hajdrihova 19, SI-1000 Ljubljana, Slovenia, School of Chemistry & Molecular Biosciences, University of Queensland, St Lucia, Brisbane, 4072 Australia, Nucleic Acid Center, Department of Physics, Chemistry and Pharmacy, University of Southern Denmark, 5230 Odense M, Denmark, EN-FIST Center of Excellence, SI-1000 Ljubljana, Slovenia and Faculty of Chemistry and Chemical Technology, University of Ljubljana, SI-1000 Ljubljana, Slovenia.

ABSTRACT
The formation of a single G-quadruplex structure adopted by a promising 25 nt G-rich vascular endothelial growth factor aptamer in a K(+) rich environment was facilitated by locked nucleic acid modifications. An unprecedented all parallel-stranded monomeric G-quadruplex with three G-quartet planes exhibits several unique structural features. Five consecutive guanine residues are all involved in G-quartet formation and occupy positions in adjacent DNA strands, which are bridged with a no-residue propeller-type loop. A two-residue D-shaped loop facilitates inclusion of an isolated guanine residue into the vacant spot within the G-quartet. The remaining two G-rich tracts of three residues each adopt parallel orientation and are linked with edgewise and propeller loops. Both 5' with 3 nt and 3' with 4 nt overhangs display well-defined conformations, with latter adopting a basket handle topology. Locked residues contribute to thermal stabilization of the adopted structure and formation of structurally pre-organized intermediates that facilitate folding into a single G-quadruplex. Understanding the impact of chemical modifications on folding, thermal stability and structural polymorphism of G-quadruplexes provides means for the improvement of vascular endothelial growth factor aptamers and advances our insights into driving nucleic acid structure by locking or unlocking the conformation of sugar moieties of nucleotides in general.

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(a) RNV66 G-quadruplex topology. Residues forming the G-quadruplex core and LNA residues are shown in orange and yellow rectangles, respectively. (b) Superimposition of the family of the 10 best structures. The two views are rotated with respect to one another to offer better overview of the structures. Guanine, LNA, adenine, thymine and cytosine residues are colored orange, yellow, red, blue and pink, respectively.
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gkt697-F4: (a) RNV66 G-quadruplex topology. Residues forming the G-quadruplex core and LNA residues are shown in orange and yellow rectangles, respectively. (b) Superimposition of the family of the 10 best structures. The two views are rotated with respect to one another to offer better overview of the structures. Guanine, LNA, adenine, thymine and cytosine residues are colored orange, yellow, red, blue and pink, respectively.

Mentions: The proposed topology of a G-quadruplex that satisfies all NMR and other spectroscopic data is parallel with four loops, and three and four residue overhangs at the 5′ and 3′ ends, respectively (Figure 4a). One of the loops, which exhibits a propeller-type topology, but with no residues involved, spans the two G-quartets and connects G6 with G7. The most unique feature of RNV66 G-quadruplex is involvement of G11 in the G-quartet formation. G7–G8 and G11 can be considered parts of two DNA strands that form the same edge of G-quadruplex core. As G8 and G11 are on the opposite ends of a three G-quartet core they are connected with a novel type of D-shaped loop consisting of two-residues T9-G10 (Figure 4a). G11 and G14 belonging to the same G-quartet are linked by a two-residue (A12-C13) edgewise loop (Figure 4a). The last propeller loop consists of two residues (C17-C18). Hydrogen-bond directions in the G-quartets are as follows: G4→G11→G14→G19, L5→G7→G15→G20 and G6→G8→G16→L21. Several NOE contacts involving residues in T1-T3 and T22-A25 segments indicate that flanking regions are reasonably well ordered. A high number of NOE correlations of residues T3 and A25 (Supplementary Figures S4 and S5) demonstrates that their position is well defined by stacking on the G4-G11-G14-G19 and G6-G8-G16-L21 quartets, respectively. NOE contacts of protons H1′, H6 and H7 of T3 with the imino protons of all four residues of the G4-G11-G14-G19 quartet (Supplementary Figure S4) affirm that these residues compose one of the outer G-quartets. It is particularly interesting for A25 as the residue at the 3′ end of the molecule to exhibit high number of NOE contacts and consequently to adopt well-defined conformation. Position of A25 leads to the formation of the T22-A25 capping structure on the G6-G8-G16-L21 quartet, resembling a basket handle.Figure 4.


G-rich VEGF aptamer with locked and unlocked nucleic acid modifications exhibits a unique G-quadruplex fold.

Marušič M, Veedu RN, Wengel J, Plavec J - Nucleic Acids Res. (2013)

(a) RNV66 G-quadruplex topology. Residues forming the G-quadruplex core and LNA residues are shown in orange and yellow rectangles, respectively. (b) Superimposition of the family of the 10 best structures. The two views are rotated with respect to one another to offer better overview of the structures. Guanine, LNA, adenine, thymine and cytosine residues are colored orange, yellow, red, blue and pink, respectively.
© Copyright Policy - creative-commons
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3814366&req=5

gkt697-F4: (a) RNV66 G-quadruplex topology. Residues forming the G-quadruplex core and LNA residues are shown in orange and yellow rectangles, respectively. (b) Superimposition of the family of the 10 best structures. The two views are rotated with respect to one another to offer better overview of the structures. Guanine, LNA, adenine, thymine and cytosine residues are colored orange, yellow, red, blue and pink, respectively.
Mentions: The proposed topology of a G-quadruplex that satisfies all NMR and other spectroscopic data is parallel with four loops, and three and four residue overhangs at the 5′ and 3′ ends, respectively (Figure 4a). One of the loops, which exhibits a propeller-type topology, but with no residues involved, spans the two G-quartets and connects G6 with G7. The most unique feature of RNV66 G-quadruplex is involvement of G11 in the G-quartet formation. G7–G8 and G11 can be considered parts of two DNA strands that form the same edge of G-quadruplex core. As G8 and G11 are on the opposite ends of a three G-quartet core they are connected with a novel type of D-shaped loop consisting of two-residues T9-G10 (Figure 4a). G11 and G14 belonging to the same G-quartet are linked by a two-residue (A12-C13) edgewise loop (Figure 4a). The last propeller loop consists of two residues (C17-C18). Hydrogen-bond directions in the G-quartets are as follows: G4→G11→G14→G19, L5→G7→G15→G20 and G6→G8→G16→L21. Several NOE contacts involving residues in T1-T3 and T22-A25 segments indicate that flanking regions are reasonably well ordered. A high number of NOE correlations of residues T3 and A25 (Supplementary Figures S4 and S5) demonstrates that their position is well defined by stacking on the G4-G11-G14-G19 and G6-G8-G16-L21 quartets, respectively. NOE contacts of protons H1′, H6 and H7 of T3 with the imino protons of all four residues of the G4-G11-G14-G19 quartet (Supplementary Figure S4) affirm that these residues compose one of the outer G-quartets. It is particularly interesting for A25 as the residue at the 3′ end of the molecule to exhibit high number of NOE contacts and consequently to adopt well-defined conformation. Position of A25 leads to the formation of the T22-A25 capping structure on the G6-G8-G16-L21 quartet, resembling a basket handle.Figure 4.

Bottom Line: Both 5' with 3 nt and 3' with 4 nt overhangs display well-defined conformations, with latter adopting a basket handle topology.Locked residues contribute to thermal stabilization of the adopted structure and formation of structurally pre-organized intermediates that facilitate folding into a single G-quadruplex.Understanding the impact of chemical modifications on folding, thermal stability and structural polymorphism of G-quadruplexes provides means for the improvement of vascular endothelial growth factor aptamers and advances our insights into driving nucleic acid structure by locking or unlocking the conformation of sugar moieties of nucleotides in general.

View Article: PubMed Central - PubMed

Affiliation: Slovenian NMR Center, National Institute of Chemistry, Hajdrihova 19, SI-1000 Ljubljana, Slovenia, School of Chemistry & Molecular Biosciences, University of Queensland, St Lucia, Brisbane, 4072 Australia, Nucleic Acid Center, Department of Physics, Chemistry and Pharmacy, University of Southern Denmark, 5230 Odense M, Denmark, EN-FIST Center of Excellence, SI-1000 Ljubljana, Slovenia and Faculty of Chemistry and Chemical Technology, University of Ljubljana, SI-1000 Ljubljana, Slovenia.

ABSTRACT
The formation of a single G-quadruplex structure adopted by a promising 25 nt G-rich vascular endothelial growth factor aptamer in a K(+) rich environment was facilitated by locked nucleic acid modifications. An unprecedented all parallel-stranded monomeric G-quadruplex with three G-quartet planes exhibits several unique structural features. Five consecutive guanine residues are all involved in G-quartet formation and occupy positions in adjacent DNA strands, which are bridged with a no-residue propeller-type loop. A two-residue D-shaped loop facilitates inclusion of an isolated guanine residue into the vacant spot within the G-quartet. The remaining two G-rich tracts of three residues each adopt parallel orientation and are linked with edgewise and propeller loops. Both 5' with 3 nt and 3' with 4 nt overhangs display well-defined conformations, with latter adopting a basket handle topology. Locked residues contribute to thermal stabilization of the adopted structure and formation of structurally pre-organized intermediates that facilitate folding into a single G-quadruplex. Understanding the impact of chemical modifications on folding, thermal stability and structural polymorphism of G-quadruplexes provides means for the improvement of vascular endothelial growth factor aptamers and advances our insights into driving nucleic acid structure by locking or unlocking the conformation of sugar moieties of nucleotides in general.

Show MeSH
Related in: MedlinePlus