Noncanonical DNA structures that stall DNA replication can cause errors in genomic DNA. Here, we investigated how the noncanonical structures formed by sequences in genes associated with a number of ...diseases impacted DNA polymerization by the Klenow fragment of DNA polymerase. Replication of a DNA sequence forming an i-motif from a telomere, hypoxia-induced transcription factor, and an insulin-linked polymorphic region was effectively inhibited. On the other hand, replication of a mixed-type G-quadruplex (G4) from a telomere was less inhibited than that of the antiparallel type or parallel type. Interestingly, the i-motif was a better inhibitor of replication than were mixed-type G4s or hairpin structures, even though all had similar thermodynamic stabilities. These results indicate that both the stability and topology of structures formed in DNA templates impact the processivity of a DNA polymerase. This suggests that i-motif formation may trigger genomic instability by stalling the replication of DNA, causing intractable diseases.
Using UV and srCD spectroscopy it is found that loop length within the i-motif structure is important for both thermal and pH stability, but in contrast to previous statements, it is the shorter ...loops that exhibit the highest stability.
I-motif formation has been confirmed in a number of gene promoter sequences known to form G-quadruplex structures. I-motif formation can occur close to physiological temperature and pH for h-tert and ...PDGF-A. The i-motif structure formed by a HIF-1α promoter sequence shows unexpected stability near neutral pH.
By using X‐ray crystallography, we show that the complexes Λ/Δ‐Ru(TAP)2(11‐CN‐dppz)2+ (TAP=1,4,5,8‐tetraazaphenanthrene, dppz=dipyridophenazine) bind DNA G‐quadruplex in an enantiospecific manner ...that parallels the specificity of these complexes with duplex DNA. The Λ complex crystallises with the normally parallel stranded d(TAGGGTTA) tetraplex to give the first such antiparallel strand assembly in which syn‐guanosine is adjacent to the complex at the 5′ end of the quadruplex core. SRCD measurements confirm that the same conformational switch occurs in solution. The Δ enantiomer, by contrast, is present in the structure but stacked at the ends of the assembly. In addition, we report the structure of Λ‐Ru(phen)2(11‐CN‐dppz)2+ bound to d(TCGGCGCCGA), a duplex‐forming sequence, and use both structural models to provide insight into the motif‐specific luminescence response of the isostructural phen analogue enantiomers.
One Way or Another: Enantiospecificity was observed in the binding of the ruthenium polypyridyl complex Λ/Δ‐Ru(TAP)2(11‐CN‐dppz)2+ to the G‐quadruplex‐forming sequence d(TAGGGTTA). Crystallographic studies yielded the first crystal structure of a ruthenium‐bound G‐quadruplex and reveal that the quadruplex adopts an antiparallel topology in the presence of the Λ isomer, but retains its parallel conformation with the Δ isomer.
To understand the molecular origins of diseases caused by ultraviolet and visible light, and also to develop photodynamic therapy, it is important to resolve the mechanism of photoinduced DNA damage. ...Damage to DNA bound to a photosensitizer molecule frequently proceeds by one-electron photo-oxidation of guanine, but the precise dynamics of this process are sensitive to the location and the orientation of the photosensitizer, which are very difficult to define in solution. To overcome this, ultrafast time-resolved infrared (TRIR) spectroscopy was performed on photoexcited ruthenium polypyridyl-DNA crystals, the atomic structure of which was determined by X-ray crystallography. By combining the X-ray and TRIR data we are able to define both the geometry of the reaction site and the rates of individual steps in a reversible photoinduced electron-transfer process. This allows us to propose an individual guanine as the reaction site and, intriguingly, reveals that the dynamics in the crystal state are quite similar to those observed in the solvent medium.
Targeting multiple malignancy features such as angiogenesis, proliferation and metastasis with one molecule is an effective strategy in developing potent anticancer agents. Ruthenium metal ...complexation to bioactive scaffolds is reported to enhance their biological activities. Herein, we evaluate the impact of Ru chelation on the pharmacological activities of two bioactive flavones (
1
and
2
) as anticancer candidates. The novel Ru complexes (
1Ru
and
2Ru
) caused a loss of their parent molecules' antiangiogenic activities in an endothelial cell tube formation assay.
1Ru
enhanced the antiproliferative and antimigratory activities of its 4-oxoflavone
1
on MCF-7 breast cancer cells (IC
50
= 66.15 ± 5 μM and 50% migration inhibition,
p
< 0.01 at 1 μM).
2Ru
diminished 4-thioflavone's (
2
) cytotoxic activity on MCF-7 and MDA-MB-231 yet significantly enhanced
2
's migration inhibition (
p
< 0.05) particularly on the MDA-MB-231 cell line. The test derivatives also showed non-intercalative interaction with VEGF and c-myc i-motif DNA sequences.
Targeting multiple malignancy features such as angiogenesis, proliferation and metastasis with one molecule is an effective strategy in developing potent anticancer agents.
The new complexes Ru(TAP)2(11‐CN‐dppz)2+, Ru(TAP)2(11‐Br‐dppz)2+and Ru(TAP)2(11,12‐diCN‐dppz)2+ are reported. The addition of nitrile substituents to the dppz ligand of the DNA photo‐oxidising ...complex Ru(TAP)2(dppz)2+ promote π‐stacking interactions and ordered binding to DNA, as shown by X‐ray crystallography. The structure of Λ‐Ru(TAP)2(11‐CN‐dppz)2+ with the DNA duplex d(TCGGCGCCGA)2 shows, for the first time with this class of complex, a closed intercalation cavity with an AT base pair at the terminus. The structure obtained is compared to that formed with the 11‐Br and 11,12‐dinitrile derivatives, highlighting the stabilization of syn guanine by this enantiomer when the terminal base pair is GC. In contrast the AT base pair has the normal Watson–Crick orientation, highlighting the difference in charge distribution between the two purine bases and the complementarity of the dppz–purine interaction. The asymmetry of the cavity highlights the importance of the purine–dppz–purine stacking interaction.
Ligand stacking: Addition of one or two terminal nitrile groups to the dppz ligand improves the stacking capability of the ligand, shown by the formation of a closed intercalation cavity.
Hydration-dependent DNA deformation has been known since Rosalind Franklin recognized that the relative humidity of the sample had to be maintained to observe a single conformation in DNA fiber ...diffraction. We now report for the first time the crystal structure, at the atomic level, of a dehydrated form of a DNA duplex and demonstrate the reversible interconversion to the hydrated form at room temperature. This system, containing d(TCGGCGCCGA) in the presence of Λ-Ru(TAP)2(dppz)2+ (TAP = 1,4,5,8-tetraazaphenanthrene, dppz = dipyrido3,2-a:2′,3′-cphenazine), undergoes a partial transition from an A/B hybrid to the A-DNA conformation, at 84–79% relative humidity. This is accompanied by an increase in kink at the central step from 22° to 51°, with a large movement of the terminal bases forming the intercalation site. This transition is reversible on rehydration. Seven data sets, collected from one crystal at room temperature, show the consequences of dehydration at near-atomic resolution. This result highlights that crystals, traditionally thought of as static systems, are still dynamic and therefore can be the subject of further experimentation.
A 3‐year prospective, randomized, placebo‐controlled trial of oral clodronate 800 mg showed that the incidence of clinical fractures was decreased by 20% in 5596 elderly women unselected for ...osteoporosis. The effect occurred in the absence of systematic calcium and vitamin D supplementation and was observed across a wide range of BMDs.
Introduction: To date, most studies with bisphosphonates have reported on their use in individuals selected to be at high risk for fracture usually by the presence of low BMD or a prior fragility fracture, usually of the spine. We wished to determine the effect of the bisphosphonate, clodronate, on the rate of fractures in women ⩾75 years of age living in the community.
Materials and Methods: Women ⩾75 years of age living in the general community in South Yorkshire and North Derbyshire, identified from general practice registers, were recruited by letter of invitation to a randomized, double‐blind, controlled trial of 800 mg oral clodronate (Bonefos) or matching placebo daily over 3 years. The main outcomes were the incidences of hip and any clinical fracture.
Results: Of the 5579 elderly women included in the intention‐to‐treat analysis of efficacy, 114 had a new hip fracture during the 3‐year treatment phase: 56 (2.0%) women in the clodronate group and 58 (2.1%) women in the placebo group (hazard ration HR, 1.02; 95% CI, 0.71–1.47). Clodronate did, however, decrease the incidence of any clinical fracture by 20% (264 women 9.5% versus 337 12.1% in the placebo group; HR, 0.80; 95% CI, 0.68–0.94). The incidence of osteoporosis‐associated nonhip fractures was also significantly decreased by 29% (5.2% versus 7.4%; HR, 0.71; 95% CI, 0.57–0.87). The ability of clodronate to reduce the risk of osteoporotic fracture was independent of baseline BMD, but the number needed‐to‐treat was lower in the presence of osteoporosis.
Conclusions: Oral daily clodronate can prevent fractures without significant adverse effects in elderly women living in the general community. The effect on hip fracture risk is not significant, but an effect similar to that at other nonvertebral sites cannot be excluded. This study suggests that antiresorptive therapies can reduce fracture incidence in high‐risk individuals even in the presence of a normal or osteopenic BMD.
Ru(phen)
(dppz)
has been studied since the 1990s due to its 'light-switch' properties. It can be used as a luminescent DNA probe, with emission switched on through DNA binding. The luminescence ...observed is dependent on the solvent accessibility of the pyrazine nitrogen atoms, and therefore is sensitive to changes in both binding site of the cation and chromophore orientation. The compound is also chiral, and there are distinct differences between the enantiomers in terms of the emission behaviour when bound to a variety of DNA sequences. Whilst a number of binary DNA-complex X-ray crystal structures are available, most include the Λ enantiomer and there is very little structural information about binding of the Δ enantiomer. Here, we present the first X-ray crystal structure of a Δ enantiomer bound to well-matched DNA, in the absence of the other, Λ enantiomer. We show how the binding site observed here can be related to a more general pattern of motifs in the crystallographic literature and propose that the Δ enantiomer can bind with five different binding modes, offering a new hypothesis for the interpretation of solution data.
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