Helix-coil transition in nucleoprotein: renaturation of polylysine-DNA and polylysine-nucleohistone complexes

Thermal denaturation and renaturation of directly mixed and reconstituted polylysine–DNA, directly mixed polylysine–nucleohistone complexes, and NaCl-treated nucleohistones in 2.5 × 10^?4 M EDTA, pH 8.0 have been studied. At the same input ratio of polylysine to DNA, the percent of renaturation of free base pairs in a directly mixed polylysine–DNA complex is higher than that in a reconstituted complex. For a directly mixed complex, the renaturation of free base pairs is proportional to the fraction of DNA bound by polylysine or inversely proportional to the sizes of free DNA loops. A of large amount of renaturation of free base pairs has also been observed for 0.6 M and 1.6 M NaCl-treated nucleohistones. The binding of polylysine to nucleohistone enhances the renaturation of histone-bound base pairs. The percent of renaturation of polylysine–bound base pairs is high and is approximately independent of the extent of binding on DNA by polylysine. This is true in polylysine–DNA complexes prepared either by reconstitution or by directly mixing. It also applies for polylysine–nucleohistone complexes. The model where polylysine-bound base pairs collapse at T_m′ with two complementary strands still bound by polylysine is favored over the model where polylysine is dissociated from DNA during melting. The low renaturation of histone-bound base pairs in nucleo-histone indicates that either histones do not hold two complementary strands of DNA tightly or that histones are fully or partially dissociated from DNA when the nucleo-histone is fully denatured.     

Theoretical study on the binding mechanism between N6-methyladenine and natural DNA bases

N6-methyladenine (m⁶A) is a rare base naturally occurring in DNA. It is different from the base adenine due to its N-CH₃. Therefore, the base not only pairs with thymine, but also with other DNA bases (cytosine, adenine and guanine). In this work, Møller-Plesset second-order (MP2) method has been used to investigate the binding mechanism between m⁶A and natural DNA bases in gas phase and in aqueous solution. The results show that N-CH₃ changed the way of N6-methyladenine binding to natural DNA bases. The binding style significantly influences the stability of base pairs. The trans-m⁶A:G and trans-m⁶A:C conformers are the most stable among all the base pairs. The existence of solvent can remarkably reduce the stability of the base pairs, and the DNA bases prefer pairing with trans-m⁶A to cis-m⁶A. Besides, the properties of these hydrogen bonds have been analyzed by atom in molecules (AIM) theory, natural bond orbital (NBO) analysis and Wiberg bond indexes (WBI). In addition, pairing with m⁶A decreases the binding energies compared to the normal Watson-Crick base pairs, it may explain the instability of the N6 site methylated DNA in theory.

Light scattering study of natural DNAs over a wide range of molecular weights: Evidence for compaction of the large molecules

This work reports light scattering measurements on DNA in aqueous solutions (100 mM NaCl, 1 mM EDTA and 10 mM Tris–HCl buffer, pH 7.8) over a wide range of molecular weights (10²–10⁵ base pairs) and shows that, in the above standard solvent, shorter chains (<10⁴ base pairs) behave as a “wormlike chain” and their diffusion coefficients as obtained by dynamic light scattering measurements, confirm the prediction of standard wormlike model, whilst longer chains (>10⁴ base pairs) behave in a different manner. Dynamic and static light scattering and SEM analysis indicate that DNA molecules 10⁵ base pairs long, condense into compact structures in our solvent conditions. Calculations done using a wormlike model are also presented and discussed in comparison both to our experimental data and to other data reported in the literature.     

A phylogenetic and biogeographic perspective on the evolution of poeciliid fishes

Phylogenetic relationships of members of the subfamily Poeciliinae (Cyprinodontiformes) are investigated to test alternate hypotheses of diversification resulting from the assembly of the Central America and the Caribbean from the Cretaceous period onwards. We use 4333 aligned base pairs of mitochondrial DNA and 1549 aligned base pairs of nuclear DNA from 55 samples representing 48 ingroup and seven outgroup species to test this hypothesis. Mitochondrial genes analyzed include those encoding the 12S and 16S ribosomal RNAs; transfer RNAs coding for valine, leucine, isoleucine, glutamine, methionine, tryptophan, alanine, asparagine, cysteine and tyrosine; and complete cytochrome b and NADH dehydrogenase subunit I and II; nuclear gene analyzed included the third exon of the recombination activation gene 1 (RAG1). Analyses of combined mtDNA and nuclear DNA data sets result in a well-supported phylogenetic hypothesis. This hypothesis is in conflict with the classical taxonomic assignment of genera into tribes and phylogenetic hypotheses based on the taxonomy; however, the molecular hypothesis defines nine clades that are geographically restricted and consistent with the geological evolution of Central America and the Caribbean. Our analyses support multiple colonization events of Middle America followed by a mix of vicariance and dispersal events.     

Isolation and some properties of DNA coding for tRNA1met from Xenopus laevis

DNA containing the reiterated genes for tRNA₁^met has been partially purified from Xenopus laevis by centrifugation in actinomycin C₁-CsCl and Ag⁺-Cs₂SO₄ gradients. These gradients separate the tRNA₁^met genes from those coding for tRNA₂^met and tRNA^val, thus confirming our earlier suggestion that these genes are not intermingled with each other (Clarkson, Birnstiel, and Purdom, 1973). The gradients also demonstrate the existence of a minor 5S DNA fraction which appears to differ from that previously isolated by Brown, Wensink, and Jordan (1971).When the enriched tDNA₁^met is digested to completion with either of the restriction endonucleases EcoRI or Hpa I, the tRNA₁^met genes are predominantly found within DNA fragments that are about 3100 base pairs long. A partial digestion with EcoRI shows that these fragments arise from the regular spacing of the enzyme restriction sites. The 3100 base pair EcoRI fragments are cleaved by Hpa I into fragments of two size classes, one of which is about 2200 base pairs long and contains the tRNA₁^met genes. The shorter fragments are about 700 base pairs long, and they appear to contain genes coding for at least one other kind of tRNA species. X. laevis tDNA₁^met thus comprises tandemly repeated DNA whose component parts show little if any length heterogeneity.     

Sensitivity of hydrogen bonds of DNA and RNA to hydration,as gauged by <Superscript>1</Superscript><Emphasis Type="Italic">J</Emphasis><Subscript>NH</Subscript> measurements in ethanol–water mixtures

Hydrogen-bond lengths of nucleic acids are (1) longer in DNA than in RNA, and (2) sequence dependent. The physicochemical basis for these variations in hydrogen-bond lengths is unknown, however. Here, the notion that hydration plays a significant role in nucleic acid hydrogen-bond lengths is tested. Watson–Crick N1...N3 hydrogen-bond lengths of several DNA and RNA duplexes are gauged using imino ¹ J _NH measurements, and ethanol is used as a cosolvent to lower water activity. We find that ¹ J _NH values of DNA and RNA become less negative with added ethanol, which suggests that mild dehydration reduces hydrogen-bond lengths even as the overall thermal stabilities of these duplexes decrease. The ¹ J _NH of DNA are increased in 8 mol% ethanol to those of RNA in water, which suggests that the greater hydration of DNA plays a significant role in its longer hydrogen bonds. The data also suggest that ethanol-induced dehydration is greater for the more hydrated G:C base pairs and thereby results in greater hydrogen-bond shortening than for the less hydrated A:T/U base pairs of DNA and RNA. Electronic Supplementary Material The online version of this article () contains supplementary material, which is available to authorized users.     

86 Characterization and differentiation of binding modes of water-soluble porphyrins at complexation with DNA

The influence of water-soluble cationic meso-tetra-(4?N-oxyethylpyridyl)porphyrin (H₂TOEPyP4) and it’s metallocomplexes with Ni, Cu, Co, and Zn on hydrodynamic and spectral behavior of DNA solutions has been studied by UV/Vis absorption and viscosity measurement. It was shown that the presence of planar porphyrins such as H₂TOEPyP4, NiTOEPyP4, and СuTOEPyP4 leads to an increase in viscosity at relatively small concentrations, and then decrease to stable values. Such behavior is explained by intercalation of these porphyrins in DNA structure because the intercalation mode involves the insertion of a planar molecule between DNA base pairs which results in a decrease in the DNA helical twist and lengthening of the DNA. Further decrease of viscosity is explained by the saturation intercalation sites and occurs outside the binding mode. But, in the case of porphyrins with axial ligands such as CoTOEPyP4 and ZnTOEPyP4, the hydrodynamic parameters decrease, which is explained by self-stacking of these porphyrins in DNA surface. This data are proved by spectral measurements. The results obtained from titration experiments were used for calculation of binding parameters: the binding constant K _b and the number of binding sites per base pair n. Obtained data reveal that K _b varies between 3.4 and 5.4?×?10⁶?M^?1 for a planar porphyrins, a range typical for intercalation mode interactions, and 5.6?×?10⁵?M^?1 and 1.8?×?10⁶?M^?1 for axial porphyrins. In addition, the exclusion parameter n also testifies that at intercalation, (n～2) the adjacent base pairs are removed to place the planar molecules, and for outside binders to pack on the surface needs too few places (n～0.5–1). It is apparent that the binding is somewhat stronger at intercalation. The viscometric and spectrophotometric measurements are in good agreement.     

Silver(I)-mediated Hoogsteen-type base pairs

Metal-mediated Hoogsteen-type base pairs are useful for the construction of DNA duplexes containing contiguous stretches of metal ions along the helical axis. To fine-tune the stability of such base pairs and the selectivity toward different metal ions, the availability of a selection of artificial nucleobases is highly desirable. In this study, we follow a theoretical approach utilizing dispersion-corrected density functional methods to evaluate a variety of artificial nucleobases as candidates for metal-mediated Hoogsteen-type base pairs. We focus on silver(I)-mediated Hoogsteen- and reverse Hoogsteen-type base pairs formed between 1-deaza- and 1,3-dideazapurine-derived nucleobases, respectively, and cytosine. Apart from two coordinative bonds, these base pairs are stabilized by a hydrogen bond. We elucidate the impact of different substituents at the C6 position and the presence or absence of an endocyclic N3 nitrogen atom on the overall stability of a base pair and concomitantly on the strength of the hydrogen and coordinative bonds. All artificial base pairs investigated in this study are less stable than the experimentally established benchmark base pair C–Ag⁺–G. The base pair formed from 1,3-dideaza-6-methoxypurine is isoenergetic to the experimentally observed C–Ag⁺–C base pair. This makes 1,3-dideaza-6-methoxypurine a promising candidate for the use as an artificial nucleobase in DNA.     

The role of the Y chromosome in human evolutionary studies

Analyses of molecular genetic data have added a new dimension to human evolutionary research. Pioneering studies of variation in human populations were based on analyses of blood groups¹ and electromorphs,² both of which represent qualitative multistate phenotypes. With the development of recombinant DNA methods in the 1970s and 1980s, the focus shifted from gene products to a new and plentiful source of human variability, restriction fragment length polymorphisms (RFLPs).^3,4 Finally, the addition of DNA sequencining survey data to the rapidly growing RFLP data base made it feasible for the first time to determine the exact number of nucleotide substitutions between different alleles, as well as to construct gene trees and reconstruct the phylogenetic history of populations.^5–7     

Interactions of DNA with divalent metal ions. IV. Competitive studies of Mn2+ binding to AT- and GC-rich DNAs

It has been inferred from previous studies that Mn²⁺ ions bind preferentially to G·C base pairs in DNA, and it has even been suggested that this preference for G·C pairs might be responsible for some of the Mn²⁺ specific effects observed in various biochemical reactions. In this paper we investigate the AT/GC preference of Mn²⁺ by direct competition studies in which AT-rich DNA was dialyzed against GC-rich DNA in the presence of varying amounts of Mn²⁺. Analysis of these results demonstrates that over a wide range of Mn²⁺/DNA(P) molar ratios, Mn²⁺ binds to A·T and to G·C base pairs with virtually identical affinity, although in a somewhat different mode. Both the present and previous nmr, uv, CD, and melting studies are discussed in terms of the different modes of binding of Mn²⁺ to single- and double-stranded DNA.     

Role of 6-Mercaptopurine in the potential therapeutic targets DNA base pairs and G-quadruplex DNA: insights from quantum chemical and molecular dynamics simulations

The theoretical studies on DNA with the anticancer drug 6-Mercaptopurine (6-MP) are investigated using theoretical methods to shed light on drug designing. Among the DNA base pairs considered, 6-MP is stacked with GC with the highest interaction energy of –46.19 kcal/mol. Structural parameters revealed that structure of the DNA base pairs is deviated from the planarity of the equilibrium position due to the formation of hydrogen bonds and stacking interactions with 6-MP. These deviations are verified through the systematic comparison between X–H bond contraction and elongation and the associated blue shift and red shift values by both NBO analysis and vibrational analysis. Bent’s rule is verified for the C–H bond contraction in the 6-MP interacted base pairs. The AIM results disclose that the higher values of electron density (ρ) and Laplacian of electron density (?²ρ) indicate the increased overlap between the orbitals that represent the strong interaction and positive values of the total electron density show the closed-shell interaction. The relative sensitivity of the chemical shift values for the DNA base pairs with 6-MP is investigated to confirm the hydrogen bond strength. Molecular dynamics simulation studies of G-quadruplex DNA d(TGGGGT)₄ with 6-MP revealed that the incorporation of 6-MP appears to cause local distortions and destabilize the G-quadruplex DNA.     

Cleavage of deoxyoxanosine-containing oligodeoxyribonucleotides by bacterial endonuclease V

Oxanine (O) is a deamination product derived from guanine with the nitrogen at the N¹ position substituted by oxygen. Cytosine, thymine, adenine, guanine as well as oxanine itself can be incorporated by Klenow Fragment to pair with oxanine in a DNA template with similar efficiency, indicating that oxanine in DNA may cause various mutations. As a nucleotide, deoxyoxanosine may substitute for deoxyguanosine to complete a primer extension reaction. Endonuclease V, an enzyme known for its enzymatic activity on uridine-, inosine- and xanthosine-containing DNA, can cleave oxanosine-containing DNA at the second phosphodiester bond 3′ to the lesion. Mg²⁺ or Mn²⁺, and to a small extent Co²⁺ or Ni²⁺, support the oxanosine-containing DNA cleavage activity. All four oxanosine-containing base pairs (A/O, T/O, C/O and G/O) were cleaved with similar efficiency. The cleavage of double-stranded oxanosine-containing DNA was ~6-fold less efficient than that of double-stranded inosine-containing DNA. Single-stranded oxanosine-containing DNA was cleaved with a lower efficiency as compared with double-stranded oxanosine-containing DNA. A metal ion enhances the binding of endonuclease V to double-stranded and single-stranded oxanosine-containing DNA 6- and 4-fold, respectively. Hypothetic models of oxanine-containing base pairs and deaminated base recognition mechanism are presented.     

Streamlining DNA Barcoding Protocols: Automated DNA Extraction and a New cox1 Primer in Arachnid Systematics

Background

DNA barcoding is a popular tool in taxonomic and phylogenetic studies, but for most animal lineages protocols for obtaining the barcoding sequences—mitochondrial cytochrome C oxidase subunit I (cox1 AKA CO1)—are not standardized. Our aim was to explore an optimal strategy for arachnids, focusing on the species-richest lineage, spiders by (1) improving an automated DNA extraction protocol, (2) testing the performance of commonly used primer combinations, and (3) developing a new cox1 primer suitable for more efficient alignment and phylogenetic analyses.

Methodology

We used exemplars of 15 species from all major spider clades, processed a range of spider tissues of varying size and quality, optimized genomic DNA extraction using the MagMAX Express magnetic particle processor—an automated high throughput DNA extraction system—and tested cox1 amplification protocols emphasizing the standard barcoding region using ten routinely employed primer pairs.

Results

The best results were obtained with the commonly used Folmer primers (LCO1490/HCO2198) that capture the standard barcode region, and with the C1-J-2183/C1-N-2776 primer pair that amplifies its extension. However, C1-J-2183 is designed too close to HCO2198 for well-interpreted, continuous sequence data, and in practice the resulting sequences from the two primer pairs rarely overlap. We therefore designed a new forward primer C1-J-2123 60 base pairs upstream of the C1-J-2183 binding site. The success rate of this new primer (93%) matched that of C1-J-2183.

Conclusions

The use of C1-J-2123 allows full, indel-free overlap of sequences obtained with the standard Folmer primers and with C1-J-2123 primer pair. Our preliminary tests suggest that in addition to spiders, C1-J-2123 will also perform in other arachnids and several other invertebrates. We provide optimal PCR protocols for these primer sets, and recommend using them for systematic efforts beyond DNA barcoding.     

Specificity of Cellular DNA-Binding Sites of Microbial Populations in a Florida Reservoir

The substrate specificity of the DNA-binding mechanism(s) of bacteria in a Florida reservoir was investigated in short- and long-term uptake studies with radiolabeled DNA and unlabeled competitors. Thymine oligonucleotides ranging in size from 2 base pairs to 19 to 24 base pairs inhibited DNA binding in 20-min incubations by 43 to 77%. Deoxynucleoside monophosphates, thymidine, and thymine had little effect on short-term DNA binding, although several of these compounds inhibited the uptake of the radiolabel from DNA in 4-h incubations. Inorganic phosphate and glucose-1-phosphate inhibited neither short- nor long-term binding of [³H]- or [³²P]DNA, indicating that DNA was not utilized as a phosphorous source in this reservoir. RNA inhibited both short- and long-term radiolabeled DNA uptake as effectively as unlabeled DNA. Collectively these results indicate that aquatic bacteria possess a generalized nucleic acid uptake/binding mechanism specific for compounds containing phosphodiester bonds and capable of recognizing oligonucleotides as short as dinucleotides. This binding site is distinct from nucleoside-, nucleotide-, phosphomonoester-, and inorganic phosphate-binding sites. Such a nucleic acid-binding mechanism may have evolved for the utilization of extracellular DNA (and perhaps RNA), which is abundant in many marine and freshwater environments.     

Molecular evolution of mitochondrial 12S RNA and cytochrome b sequences in the pantherine lineage of Felidae

DNA sequence comparisons of two mitochondrial DNA genes were used to infer phylogenetic relationships among 17 Felidae species, notably 15 in the previously described pantherine lineage. The polymerase chain reaction (PCR) was used to generate sequences of 358 base pairs of the mitochondrial 12S RNA gene and 289 base pairs of the cytochrome b protein coding gene. DNA sequences were compared within and between 17 felid and five nonfelid carnivore species. Evolutionary trees were constructed using phenetic, cladistic, and maximum likelihood algorithms. The combined results suggested several phylogenetic relationships including (1) the recognition of a recently evolved monophyletic genus Panthera consisting of Panthera leo, P. pardus, P. onca, P. uncia, P. tigris, and Neofelis nebulosa; (2) the recent common ancestry of Acinonyx jubatus, the African cheetah, and Puma concolor, the American puma; and (3) two golden cat species, Profelis temmincki and Profelis aurata, are not sister species, and the latter is strongly associated with Caracal caracal. These data add to the growing database of vertebrate mtDNA sequences and, given the relatively recent divergence among the felids represented here (1-10 Myr), allow 12S and cytochrome b sequence evolution to be addressed over a time scale different from those addressed in most work on vertebrate mtDNA.      

An intercalation-locked parallel-stranded DNA tetraplex

DNA has proved to be an excellent material for nanoscale construction because complementary DNA duplexes are programmable and structurally predictable. However, in the absence of Watson–Crick pairings, DNA can be structurally more diverse. Here, we describe the crystal structures of d(ACTCGGATGAT) and the brominated derivative, d(AC^BrUCGGA^BrUGAT). These oligonucleotides form parallel-stranded duplexes with a crystallographically equivalent strand, resulting in the first examples of DNA crystal structures that contains four different symmetric homo base pairs. Two of the parallel-stranded duplexes are coaxially stacked in opposite directions and locked together to form a tetraplex through intercalation of the 5′-most A–A base pairs between adjacent G–G pairs in the partner duplex. The intercalation region is a new type of DNA tertiary structural motif with similarities to the i-motif. ¹H–¹H nuclear magnetic resonance and native gel electrophoresis confirmed the formation of a parallel-stranded duplex in solution. Finally, we modified specific nucleotide positions and added d(GAY) motifs to oligonucleotides and were readily able to obtain similar crystals. This suggests that this parallel-stranded DNA structure may be useful in the rational design of DNA crystals and nanostructures.     

Repetitive DNA in Yeasts

BETWEEN 10% and 70% of the nuclear DNA of all higher organisms consists of repeating sequences^1,2 (in some organisms only 6–13 base pairs long³) which comprise families of identical or similar base sequences repeated from several hundred to more than a million times. Much of this is not transcribed⁴ and the most repetitive sequences are located in the centromeric heterochromatin⁵. If repetitive DNA occurs in all eukaryotic cells, however, it is surprising that in renaturation studies it has not been found in yeast^2,6. In Saccharomyces cerevisiae,a large number of the AT base pairs of the mitochondrial DNA probably occur in poly AT sequences^7,8. This may result in unusual renaturation kinetics.     

Characterization and use of an unprecedentedly bright and structurally non-perturbing fluorescent DNA base analogue

This article presents the first evidence that the DNA base analogue 1,3-diaza-2-oxophenoxazine, tC^O, is highly fluorescent, both as free nucleoside and incorporated in an arbitrary DNA structure. tC^O is thoroughly characterized with respect to its photophysical properties and structural performance in single- and double-stranded oligonucleotides. The lowest energy absorption band at 360 nm (ε = 9000 M⁻¹ cm⁻¹) is dominated by a single in-plane polarized electronic transition and the fluorescence, centred at 465 nm, has a quantum yield of 0.3. When incorporated into double-stranded DNA, tC^O shows only minor variations in fluorescence intensity and lifetime with neighbouring bases, and the average quantum yield is 0.22. These features make tC^O, on average, the brightest DNA-incorporated base analogue so far reported. Furthermore, it base pairs exclusively with guanine and causes minimal perturbations to the native structure of DNA. These properties make tC^O a promising base analogue that is perfectly suited for e.g. photophysical studies of DNA interacting with macromolecules (proteins) or for determining size and shape of DNA tertiary structures using techniques such as fluorescence anisotropy and fluorescence resonance energy transfer (FRET).     

Reassociation Kinetics and Cytophotometric Characterization of Peanut (Arachis hypogaea L.) DNA

The base composition of peanut (var. NC-17) DNA determined from thermal denaturation profiles showed an average guanine plus cystosine content of 34% which was in close approximation to 36% guanine plus cytosine calculated from the buoyant density. Buoyant density also indicated the absence of satellite DNA. The genome size, 2.0 × 10⁹ base pairs, as determined by reassociation kinetics of the single copy DNA was close to the genome size determined by cytophotometry, 2.1 × 10⁹ base pairs. Peanut DNA averaging 450 to 600 base pairs long, reassociated in phosphate buffer and fractionated by hydroxylapatite, indicated a DNA genome composition of 36% nonrepetitive or single copy DNA; reassociation in formamide and followed by optical methods indicated the repetitive DNA possesses highly repeated, intermediately repeated and rarely repeated components of DNA with DNA sequences repeated on the average about 38,000, 6,700, and 200 times each. Different criteria of reassociation in formamide revealed further subdivisions of these four separate components of DNA. The DNA of above mentioned NC-17 variety compared to Florigiant variety showed no differences in thermal denaturation profiles, buoyant density, or in genome size.     

Imino 1H- and 31P-NMR analysis of the interaction of propidium and ethidium with DNA

At low temperature and low salt concentration, both imino proton and ³¹p-nmr spectra of DNA complexes with the intercalators ethidium and propidium are in the slow-exchange region. Increasing temperature and/or increasing salt concentration results in an increase in the site exchange rate. Ring-current effects from the intercalated phenanthridinium ring of ethidium and propidium cause upfield shifts of the imino protons of A · T and G · C base pairs, which are quite similar for the two intercalators. The limiting induced chemical shifts for propidium and ethidium at saturation of DNA binding sites are approximately 0.9 ppm for A · T and 1.1 ppm for G · C base pairs. The similarity of the shifts for ethidium and propidium, in both the slow- and fast-exchange regions over the entire titration of DNA, shows that a binding model for propidium with neighbor-exclusion binding and negative ligand cooperativity is correct. The fact that a unique chemical shift is obtained for imino protons at intercalated sites over the entire titration and that no unshifted imino proton peaks remain at saturation binding of ethidium and propidium supports a neighbor-exclusion binding model with intercalators bound at alternating sites rather than in clusters on the double helix. Addition of ethidium and propidium to DNA results in downfield shifts in ³¹P-nmr spectra. At saturation ratios of intercalator to DNA base pairs in the titration, a downfield shoulder (approximately ?2.7 ppm) is apparent, which accounts for approximately 15% of the spectral area. The main peak is at ?3.9 to ?4.0 ppm relative to ?4.35 in uncomplexed DNA. The simplest neighbor-binding model predicts a downfield peak with approximately 50% of the spectral area and an upfield peak, near the chemical shift for uncomplexed DNA, with 50% of the area. This is definitely not the case with these intercalators. The observed chemical shifts and areas for the DNA complexes can be explained by models, for example, that involve spreading the intercalation-induced unwinding of the double helix over several base pairs and/or a DNA sequence- and conformation-dependent heterogeneity in intercalation-induced chemical shifts and resulting exchange rates.