Left-handed Z-form Research Articles

Chelerythrine, a new benzo[c]phenanthridine based converter of alternate DNA structure into conventional one: A spectroscopic exploration

Association of different polymprphic forms of DNA having alternating G-C sequences with the alkaloid Chelerythrine Chloride (CHL) was probed by using UV absorption and circular dichroism (CD) spectrometry. Spectrophotometric titration revealed that the association of CHL with right handed B-form was non-cooperative in nature with higher binding affinity (∼106 M−1) in contrast to the cooperative binding with left handed Z-form and HL-form where the intrinsic binding constants were of the order of ∼104 M−1. Further, conversion of the left-handed Z-form and the HL-form of the polymer back to the bound right-handed form by CHL has been evidenced from CD spectroscopy and under our experimental condition, the number of base pairs of Z-form and HL-form that switched to right-handed conformation by each bound CHL molecule was found to be 2–3. Moreover, kinetic measurements showed that in presence of increasing concentration of CHL, both the rate and the extent of B-form to Z-form transition were reduced.

Permanganate/S1 Nuclease Footprinting Reveals Non-B DNA Structures with Regulatory Potential across a Mammalian Genome

DNA in cells is predominantly B-form double helix. Though certain DNA sequences invitro may fold into other structures, such as triplex, left-handed Zform, or quadruplex DNA, the stability and prevalence of these structures invivo are not known. Here, using computational analysis of sequence motifs, RNA polymerase II binding data, and genome-wide potassium permanganate-dependent nuclease footprinting data, we map thousands of putative non-B DNA sites at high resolution in mouse B cells. Computational analysis associates these non-B DNAs with particular structures and indicates that they form at locations compatible with an involvement in gene regulation. Further analyses support the notion that non-B DNA structure formation influences the occupancy and positioning of nucleosomes in chromatin. These results suggest that non-B DNAs contribute to the control of a variety of critical cellular and organismal processes.

FACT is a sensor of DNA torsional stress in eukaryotic cells.

Transitions of B-DNA to alternative DNA structures (ADS) can be triggered by negative torsional strain, which occurs during replication and transcription, and may lead to genomic instability. However, how ADS are recognized in cells is unclear. We found that the binding of candidate anticancer drug, curaxin, to cellular DNA results in uncoiling of nucleosomal DNA, accumulation of negative supercoiling and conversion of multiple regions of genomic DNA into left-handed Z-form. Histone chaperone FACT binds rapidly to the same regions via the SSRP1 subunit in curaxin-treated cells. In vitro binding of purified SSRP1 or its isolated CID domain to a methylated DNA fragment containing alternating purine/pyrimidines, which is prone to Z-DNA transition, is much stronger than to other types of DNA. We propose that FACT can recognize and bind Z-DNA or DNA in transition from a B to Z form. Binding of FACT to these genomic regions triggers a p53 response. Furthermore, FACT has been shown to bind to other types of ADS through a different structural domain, which also leads to p53 activation. Thus, we propose that FACT acts as a sensor of ADS formation in cells. Recognition of ADS by FACT followed by a p53 response may explain the role of FACT in DNA damage prevention.

Carbon Nanomaterials and DNA: from Molecular Recognition to Applications.

DNA is polymorphic. Increasing evidence has indicated that many biologically important processes are related to DNA's conformational transition and assembly states. In particular, noncanonical DNA structures, such as the right-handed A-form, the left-handed Z-form, the triplex, the G-quadruplex, the i-motif, and so forth, have been specific targets for the diagnosis and therapy of human diseases. Meanwhile, they have been widely used in the construction of smart DNA nanomaterials and nanoarchitectures. As rising stars in materials science, the family of carbon nanomaterials (CNMs), including two-dimensional graphene, one-dimensional carbon nanotubes (CNTs), and zero-dimensional graphene or carbon quantum dots (GQDs or CQDs), interact with DNA and are able to regulate the conformational transitions of DNA. The interaction of DNA with CNMs not only opens new opportunities for specific molecular recognition, but it also expands the promising applications of CNMs from materials science to biotechnology and biomedicine. In this Account, we focus on our contributions to the field of interactions between CNMs and DNA in which we have explored their promising applications in nanodevices, sensing, materials synthesis, and biomedicine. For one-dimensional CNTs, two-dimensional graphene, and zero-dimensional GQDs and CQDs, the basic principles, binding modes, and applications of the interactions between CNMs and DNA are reviewed. We aim to give prominence to the important status of CNMs in the field of molecular recognition for DNA. First, we summarized our discovery of the interactions between single-walled carbon nanotubes (SWNTs) with duplex, triplex, and human telomeric i-motif DNA and their interesting applications. For example, SWNTs are the first chemical agents that can selectively stabilize human telomeric i-motif DNA and induce its formation under physiological conditions. On the basis of this principle, two types of nanodevices were designed. One was used for highly sensitive detection of ppm levels of SWNTs in cells, and the other monitored i-motif DNA formation. Further studies indicated that SWNTs could inhibit telomerase activity in living cells and cause telomere dysfunction, providing new insight into the biological effects of SWNTs. Then, some applications that are based on the interactions between graphene and DNA are also summarized. Combined with other nanomaterials, such as metal and upconversion nanoparticles, several hybrid nanomaterials were successfully constructed, and a series of DNA logic gates were successfully developed. Afterwards, the newcomer of the carbon nanomaterials family, carbon quantum dots (CQDs), were found to be capable of modulating right-handed B-form DNA to left-handed Z-form DNA. These were further used to design FRET logic gates that were based on the CQD-derived DNA conformational transition. Taking into account the remaining challenges and promising aspects, CNM-based DNA nanotechnology and its biomedical applications will attract more attention and produce new breakthroughs in the near future.

Formation of Poly[d(A-T)2] Specific Z-DNA by a Cationic Porphyrin.

Typical CD spectrum of the right-handed poly[d(A-T)2] was reversed when trans-bis(N-methylpyrimidium-4-yl)diphenyl porphyrin (trans-BMPyP) was bound, suggesting that the helicity of the polynucleotide was reversed to the left-handed form. The formation of the left-handed Z-form poly[d(A-T)2] was confirmed by 31P NMR, in which a single 31P peak of B-form poly[d(A-T)2] was split into two peaks, which is similar to the conventional B-Z transition of poly[d(G-C)2] induced by the high ionic strength. The observed B-Z transition is unique for poly[d(A-T)2]. The other polynucleotides, including poly[d(G-C)2], poly(dG)·poly(dC) and poly(dA)·poly(dT) remained as the right-handed form in the presence of the same porphyrin. This observation suggests that the porphyrin array that was formed along the poly[d(A-T)2] provides a template to which left-handed poly[d(A-T)2] is associated with an electrostatic interaction.

Annealed Random Copolymer Model of the B-Z Transition in DNA: Torsional Responses

Both in vivo and in vitro, specific sequences in double-stranded DNA can adopt the left-handed Z-form when underwound. Recently, the B-Z transition of DNA has been studied in detail in magnetic tweezers experiments by several groups. We present a theoretical description of this transition, based on an annealed random copolymer model. The transition of a switchable sequence is discussed as a function of energetic and geometric parameters of the B- and Z-forms, of the applied boundary conditions, and of the characteristics of the B-Z interface. We address a possible torsional softening upon the B-Z transition. The model can be also applied to other biofilaments with annealed torsional/flexural degrees of freedom.

Chiroptical properties of anionic and cationic porphyrins and metalloporphyrins in complex with left-handed Z-DNA and right-handed B-DNA

We report the chiroptical signature and binding interactions of cationic (meso-tetrakis(4-N-methylptridyl)porphyrin, 2HT4) and anionic (meso-tetrakis(4-sulfonatophenyl)porphyrin, 2HTPPS) porphyrins and their zinc(II) and nickel(II) derivatives (ZnT4, ZnTPPS, NiT4, and NiTPPS) with right-handed B-form and two forms of left-handed Z-form of alternating guanine–cytosine polydeoxynucleotide poly(dG–dC)2. NiTPPS is able to spectroscopically discriminate between spermine-induced Z-DNA and Co(III)-induced Z-DNA via new induced circular dichroism signal in the visible region of the electromagnetic spectrum.

Vibrations of ordered counterions around left- and right-handed DNA double helixes

The vibrations of ordered counterions around right- and left-handed DNA double helix are studied. To determine the modes of DNA conformational vibrations the structure of the double helix with counterions is considered as ionic lattice (ion-phosphate lattice). Using the developed approach the frequencies and Raman intensities for right-handed B-form and left-handed Z-form of the double helix with Na+, K+, Rb+, Cs+, and Mg2+ counterions are calculated. The obtained frequencies of vibrations of internal structure elements of the double helix (<100 cm−1) weakly depend on counterion type. In contrast, the vibrations of the ion-phosphate lattice are determined by counterion mass and charge. The frequencies of ion-phosphate vibrations of alkali metal counterions decrease from 180 to 100 cm−1, while their Raman intensities increase as the counterion mass increases for the both B- and Z-DNA. In the case of Z-DNA new mode of ion-phosphate vibrations near 150 cm−1 is found. This mode is characterized by vibrations of Mg2+ counterions with respect to the phosphates of different strands of the double helix. Our results explain the experimental Raman spectra of Z-DNA.

ChemInform Abstract: Conformational Changes of Non‐B DNA

AbstractReview: 189 refs.

Quinacrine and 9-Amino Acridine Inhibit B-Z and B-HL Form DNA Conformational Transitions

The interaction of quinacrine and 9-amino acridine with right-handed B-form, left-handed Z-form, and left-handed protonated (H(L))-form structures of polydG-me(5)dC was investigated by circular dichroism and absorption spectral analysis. Both the compounds bind strongly to the B-form structure and convert the Z-form and H(L)-form back to the bound right-handed form. Circular dichroic data revealed that the conformation at the binding site is right-handed even though adjacent regions of the polynucleotide may have left-handed conformation. The rate and extent of B-form-to-Z-form transition were decreased in the presence of these compounds. Scatchard analysis revealed that both quinacrine and 9-amino acridine bind strongly to the polynucleotide in the B-form in a noncooperative manner, in sharp contrast to the highly cooperative binding to the Z-form and H(L)-form. Results indicated that the cooperative binding of these drugs with the Z-form and the H(L)-forms was associated with a sequential conversion of the polynucleotide from a left-handed to a bound right-handed conformation. Experimental data enabled the calculation of the number of base pairs of Z-form (7-8 with quinacrine and 9-amino acridine) and H(L)-form (4 and 25, respectively, with quinacrine and 9-amino acridine) that adopt a right-handed conformation for each bound ligand. As these compounds are known to bind preferentially to alternating guanine--cytosine sequences, which are capable of easily undergoing the B-to-Z or B-to-H(L) transition, these effects may be important in understanding their biological activities.

Conformational changes of non-B DNA

In contrast to B-DNA that has a right-handed double helical structure with Watson-Crick base pairing under the ordinary physiological conditions, repetitive DNA sequences under certain conditions have the potential to fold into non-B DNA structures such as hairpin, triplex, cruciform, left-handed Z-form, tetraplex, A-motif, etc. Since the non-B DNA-forming sequences induce the genetic instability and consequently can cause human diseases, the molecular mechanism for their genetic instability has been extensively investigated. On the contrary, non-B DNA can be widely used for application in biotechnology because many DNA breakage hotspots are mapped in or near the sequences that have the potential to adopt non-B DNA structures. In addition, they are regarded as a fascinating material for the nanotechnology using non-B DNAs because they do not produce any toxic byproducts and are robust enough for the repetitive working cycle. This being the case, an understanding on the mechanism and dynamics of their structural changes is important. In this critical review, we describe the latest studies on the conformational dynamics of non-B DNAs, with a focus on G-quadruplex, i-motif, Z-DNA, A-motif, hairpin and triplex (189 references).

Can Copper(II) Mediate Hoogsteen Base‐Pairing in a Left‐Handed DNA Duplex? A Pulse EPR Study

Pulse EPR spectroscopy is used to investigate possible structural features of the copper(II) ion coordinated to poly(dG-dC).poly(dG-dC) in a frozen aqueous solution, and the structural changes of the polynucleotide induced by the presence of the metal ion. Two different copper species were identified and their geometry explained by a molecular model. According to this model, one species is exclusively coordinated to a single guanine with the N7 nitrogen atom forming a coordinative bond with the copper. In the other species, a guanine and a cytosine form a ternary complex together with the copper ion. A copper crosslink between the N7 of guanine and N3 of cytosine is proposed as the most probable coordination site. Moreover, no evidence was found for an interaction of either copper species with a phosphate group or equatorial water molecules. In addition, circular dichroism (CD) spectroscopy showed that the DNA of the Cu(II)-poly(dG-dC).poly(dG-dC) adducts resembles the left-handed Z-form. These results suggest that metal-mediated Hoogsteen base pairing, as previously proposed for a right-handed DNA duplex, can also occur in a double-stranded left-handed DNA.

Interactions of a Tetraanionic Porphyrin with DNA: from a Z-DNA Sensor to a Versatile Supramolecular Device

The anionic nickel(II) porphyrin NiTPPS is able to selectively sense the spermine induced left-handed Z-form of DNA while it is completely silent in the presence of right-handed B-DNA. Interactions between the DNA and the porphyrin can be easily modulated by pH and temperature. The resulting Z-DNA-porphyrin-spermine complex behaves as a supramolecular reversible information storage system and as a reversible AND logic gate.

Interaction of rac-[Cu(diimine)3]2+ and rac-[Zn(diimine)3]2+ complexes with CT DNA: effect of fluxional Cu(ii) geometry on DNA binding, ligand–promoted exciton coupling and prominent DNA cleavage

The complexes [Cu(phen)(3)](ClO(4))(2) 1, [Cu(5,6-dmp)(3)](ClO(4))(2) 2, [Cu(dpq)(3)](ClO(4))(2) 3, [Zn(phen)(3)](ClO(4))(2) 4, [Zn(5,6-dmp)(3)](ClO(4))(2) 5 and [Zn(dpq)(3)](ClO(4))(2) 6, where phen = 1,10-phenanthroline, 5,6-dmp = 5,6-dimethyl-1,10-phenanthroline and dpq = dipyrido[3,2-d:2',3'-f]quinoxaline, have been isolated, characterized and their interaction with calf thymus DNA studied by using a host of physical methods. The X-ray crystal structures of rac-[Cu(5,6-dmp)(3)](ClO(4))(2) and rac-[Zn(5,6-dmp)(3)](ClO(4))(2) have been determined. While 2 possesses a regular elongated octahedral coordination geometry (REO), 5 possesses a distorted octahedral geometry. Absorption spectral titrations of the Cu(II) complexes with CT DNA reveal that the red-shift (12 nm) and DNA binding affinity of 3 (K(b), 7.5 x 10(4) M(-1)) are higher than those of 1 (red-shift, 6 nm; K(b), 9.6 x 10(3) M(-1)) indicating that the partial insertion of the extended phen ring of dpq ligand in between the DNA base pairs is deeper than that of phen ring. Also, 2 with a fluxional Cu(II) geometry interacts with DNA (K(b), 3.8 x 10(4) M(-1)) more strongly than 1 suggesting that the hydrophobic forces of interaction of 5,6 methyl groups on the phen ring is more pronounced than the partial intercalation of phen ring in the latter with a static geometry. The DNA binding affinity of 1 is lower than that of its Zn(ii) analogue 4, and, interestingly, the DNA binding affinity 2 of with a fluxional geometry is higher than that of its Zn(II) analogue 5 with a spherical geometry. It is remarkable that upon binding to DNA 3 shows an increase in viscosity higher than that the intercalator EthBr does, which is consistent with the above DNA binding affinities. The CD spectra show only one induced CD band on the characteristic positive band of CT DNA upon interaction with the phen (1,4) and dpq (3,6) complexes. In contrast, the 5,6-dmp complexes 2 and 5 bound to CT DNA show exciton-coupled biphasic CD signals with 2 showing CD signals more intense than 5. The Delta-enantiomer of rac-[Cu(5,6-dmp)(3)](2+) 2 binds specifically to the right-handed B-form of CT DNA at lower ionic strength (0.05 M NaCl) while the Lambda-enantiomer binds specifically to the left-handed Z-form of CT DNA generated by treating the B-form with 5 M NaCl. The complex 2 is stabilized in the higher oxidation state of Cu(II) more than its phen analogue 1 upon binding to DNA suggesting the involvement of electrostatic forces in DNA interaction of the former. In contrast, 3 bound to DNA is stabilized as Cu(I) rather than the Cu(II) oxidation state due to partial intercalative interaction of the dpq ligand. The efficiencies of the complexes to oxidatively cleave pUC19 DNA vary in the order, 3> 1 >> 2 with 3 effecting 100% cleavage even at 10 microM complex concentration. However, interestingly, this order is reversed when the DNA cleavage is performed using H(2)O(2) as an activator and the highest cleavage efficiency of 2 is ascribed to its electrostatic interaction with the exterior phosphates of DNA.

Interaction between the Z-Type DNA Duplex and 1,3-Propanediamine: Crystal Structure of d(CACGTG)2 at 1.2 Å Resolution,

The crystal structure of a hexamer duplex d(CACGTG)(2) has been determined and refined to an R-factor of 18.3% using X-ray data up to 1.2 A resolution. The sequence crystallizes as a left-handed Z-form double helix with Watson-Crick base pairing. There is one hexamer duplex, a spermine molecule, 71 water molecules, and an unexpected diamine (Z-5, 1,3-propanediamine, C(3)H(10)N(2)) in the asymmetric unit. This is the high-resolution non-disordered structure of a Z-DNA hexamer containing two AT base pairs in the interior of a duplex with no modifications such as bromination or methylation on cytosine bases. This structure does not possess multivalent cations such as cobalt hexaammine that are known to stabilize Z-DNA. The overall duplex structure and its crystal interactions are similar to those of the pure-spermine form of the d(CGCGCG)(2) structure. The spine of hydration in the minor groove is intact except in the vicinity of the T5A8 base pair. The binding of the Z-5 molecule in the minor grove of the d(CACGTG)(2) duplex appears to have a profound effect in conferring stability to a Z-DNA conformation via electrostatic complementarity and hydrogen bonding interactions. The successive base stacking geometry in d(CACGTG)(2) is similar to the corresponding steps in d(CG)(3). These results suggest that specific polyamines such as Z-5 could serve as powerful inducers of Z-type conformation in unmodified DNA sequences with AT base pairs. This structure provides a molecular basis for stabilizing AT base pairs incorporated into an alternating d(CG) sequence.

Interaction of rac-[Ru(5,6-dmp)3]2+ with DNA: Enantiospecific DNA Binding and Ligand-Promoted Exciton Coupling

The X-ray crystal structure of the complex rac-[Ru(5,6-dmp)(3)]Cl(2) (5,6-dmp = 5,6-dimethyl-1,10-phenanthroline) reveals a distorted octahedral coordination geometry with the Ru-N bond distances shorter than in its phen analogue. Absorption spectral titrations with CT DNA reveal that rac-[Ru(5,6-dmp)(3)](2+) interacts (K(b), (8.0 +/- 0.2) x 10(4) M(-1)) much more strongly than its phen analogue. The emission intensity of the 5,6-dmp complex is dramatically enhanced on binding to DNA, which is higher than that of the phen analogue. Also, interestingly, time-resolved emission measurements on the DNA-bound complex shows biexponential decay of the excited states with the lifetimes of short- and long-lived components being higher than those for the phen analogue. The CD spectral studies of rac-[Ru(5,6-dmp)(3)](2+) bound to CT DNA provide a definite and elegant evidence for the enantiospecific interaction of the complex with B-form DNA. Competitive DNA binding studies using rac-[Ru(phen)(3)](2+) provide support for the strong binding of the complex with DNA. The Delta-enantiomer of rac-[Ru(5,6-dmp)(3)](2+) binds specifically to the right-handed B-form of poly d(GC)(12) at lower ionic strength (0.05 M NaCl), and the Lambda-enantiomer binds specifically to the left-handed Z-form of poly d(GC)(12) generated by treating the B-form with 5 M NaCl. The strong electronic coupling of the DNA-bound complex with the unbound complex facilitates the change in its enantiospecificity upon changing the conformation of DNA. The (1)H NMR spectra of rac-[Ru(5,6-dmp)(3)](2+) bound to poly d(GC)(12) reveal that the complex closely interacts most possibly in the major grooves of DNA. Electrochemical studies using ITO electrode show that the 5,6-dmp complex stabilizes CT DNA from electrocatalytic oxidation of its guanine base more than the phen analogue does.

Structure of d(TGCGCG)·d(CGCGCA) in two crystal forms: effect of sequence and crystal packing in Z-DNA

The sequence d(TGCGCG).d(CGCGCA) crystallized in two crystal forms, orthorhombic and hexagonal, in the presence of cobalt hexammine chloride, a known inducer of the left-handed Z-form of DNA. The crystal structures have been solved and refined at 1.71 A resolution in space group P2(1)2(1)2(1) and 2.0 A resolution in space group P6(5). The orthorhombic structure contains one Z-DNA hexamer duplex, while the hexagonal structure contains two hexamer duplexes in the structure. Of the latter, one is situated on a crystallographic sixfold screw axis, leading to disorder. This paper reports the effects of sequence and crystal packing on the structure of Z-type DNA. The structures lend additional support to the authors' earlier conclusion that a stretch of four C.G base pairs is sufficient to nucleate and define the regular model of the left-handed helix based on the structure of d(CGCGCG)2.

Circular Dichroism and NMR Studies of Metabolically Stableα-Methylpolyamines: Spectral Comparison with Naturally Occurring Polyamines

Three synthetic polyamine analogs, α-methylspermine, and α,α′-dimethylspermine, were compared with their naturally occurring counterparts, spermidine and spermine, by two different spectral techniques. The interaction of polyamines with oligodeoxynucleotides was measured by circular dichroism in order to monitor the polyamine-induced conversion of right-handed B-DNA to the left-handed Z-form. The methylated analogs were shown to be equally effective as the natural polyamines in inducing the B → Z transition. The pH dependence of the chemical shift of all carbon atoms in each of the five polyamines was measured by 13C-NMR spectroscopy. With the exception of expected changes in chemical shift due to the presence of the α-methyl substituents, the chemical shifts and pH dependence of all carbon atoms in the three α-methyl polyamines were similar to the corresponding naturally occurring polyamines. The combined data indicate that α-methyl polyamines have physical properties that are very similar to their natural counterparts. The two metabolically stable polyamine analogs, α-methylspermidine and α,α′-dimethylspermine, are therefore useful surrogates for spermidine and spermine in the study of numerous polyamine-mediated effects in mammalian cell cultures and can be used in such studies without the requirement for coadministration of amine oxidase inhibitors.

Conformation Change of Poly(dG-dC)•Poly(dG-dC) in Cationic Polyamine Liposome Complexes: Effect of Charge Density and Flexibility of Amine Chains in Headgroups

Identification of DNA conformations in delivery cargoes is important to understand the transfection mechanisms. Conformations of poly(dG-dC)•poly(dG-dC), as a model of DNA, were studied in two different cationic liposome complexes prepared from N-[1-(2,3-dioleyloxy)propyl]-N,N,N-trimethylammonium chloride (DOTMA) and N,NI,NII,NIII-tetramethyl-N,NI,NII,NIII-tetrapalmithyl spermine (TM-TPS) with a helper lipid, dioleoyl phosphotidylethanolamine (DOPE) by Raman microscopy. Characteristic spectral changes in guanine peaks and cytosine peaks of the poly(dG-dC)•poly(dG-dC) distinguished between the right-handed B-form and left-handed Z-form inside the cationic liposome complexes. The poly(dG-dC)•poly(dG-dC) in the TM-TPS/DOPE liposome undergoes the B−Z transition while no conformation change was observed in the poly(dG-dC)•poly(dG-dC) inside the DOTMA/DOPE liposome. Each multi-charged TM-TPS molecule contains four cationic nitrogen atoms in the headgroup. The DOTMA molecule with less cationic nitrogens does not...

Structure of d(CACGCG).d(CGCGTG) in crystals grown in the presence of ruthenium III hexammine chloride.

Hexammine ions are strong inducers of the transition from the B-form to the left-handed Z-form in DNA. Here the structure of d(CACGCG). d(CGCGTG) obtained from crystals grown from a drop containing [Ru(NH3)6]Cl3 is reported. The structure is clearly characterized as Z-DNA. When compared with the structure of d(CACGCG).d(CGCGTG)/MgCl2 and that of d(CGCGCG)2, subtle differences are seen, most noticeably in the water structure. Since stable well diffracting crystals grow easily in the presence of [Ru(NH3)6]Cl3 and since this ion is not visible in the electron density it is concluded that the ion plays a non-specific role in stabilizing Z-DNA.