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PubMed1992

Role of magnesium ion in the interaction between chromomycin A3 and DNA: binding of chromomycin A3-Mg2+ complexes with DNA.

Chromomycin A3 is an antitumor antibiotic which blocks macromolecular synthesis via reversible interaction with DNA template only in the presence of divalent metal ions such as Mg2+. The role of Mg2+ in this antibiotic-DNA interaction is not well understood. We approached the problem in two steps via studies on the interaction of (i) chromomycin A3 and Mg2+ and (ii) chromomycin A3-Mg2+ complex(es) and DNA. Spectroscopic techniques such as absorption, fluorescence, and CD were employed for this purpose. The results could be summed up in two parts. Absorption, fluorescence, and CD spectra of the antibiotic change upon addition of Mg2+ due to complex formation between them. Analysis of the quantitative dependence of change in absorbance of chromomycin A3 (at 440 nm) upon input concentration of Mg2+ indicates formation of two types of complexes with different stoichiometries and formation constants. Trends in change of fluorescence and CD spectroscopic features of the antibiotic in the presence of Mg2+ at different concentrations further corroborate this result. The two complexes are referred to as complex I (with 1:1 stoichiometry in terms of chromomycin A3:Mg2+) and complex II (with 2:1 stoichiometry in terms of chromomycin A3:Mg2+), respectively, in future discussions. The interactions of these complexes with calf thymus DNA were examined to check whether they bind differently to the same DNA. Evaluation of binding parameters, intrinsic binding constants, and binding stoichiometry, by means of spectrophotometric and fluorescence titrations, shows that they are different. Distinctive spectroscopic features of complexes I and II, when they are bound to DNA, also support that they bind differently to the above DNA. Measurement of thermodynamic parameters characterizing their interactions with calf thymus DNA shows that complex I-DNA interaction is exothermic, in contrast to complex II-DNA interaction, which is endothermic. This feature implies a difference in the molecular nature of the interactions between the complexes and calf thymus DNA. These observations are novel and significant to understand the antitumor property of the antibiotic. They are also discussed to provide explanations for the earlier reports that in some cases appeared to be contradictory.

Animals↗

PubMed1992

Interaction between antitumor antibiotic chromomycin A3 and Mg2+. I. Evidence for the formation of two types of chromomycin A3-Mg2+ complexes.

Chromomycin A3 (CHRA3) is an antitumor antibiotic which binds to Mg2+. In the present communication, we show, by means of equilibrium spectroscopic studies (such as absorption, fluorescence and circular dichroism), that two types of CHRA3-Mg2+ complexes (of 1:1 and 1.9:1 stoichiometries in terms of CHRA3:Mg2+, respectively) are formed depending on the concentrations of CHRA3 and Mg2+. The rate constant and activation energy for the formation of two complexes are different, thereby reinforcing the proposition that they are different molecular species. This observation is novel and significant in order to understand the anticancer property of the drug. It also provides explanation for earlier observations that site, affinity parameters and mode of interaction of the drug with DNA in the presence of Mg2+ depend on the relative concentration of Mg2+.

Chromomycin A3↗

PubMed1985

NMR studies of chromomycin A3 interaction with DNA.

The binding of chromomycin A3 to calf thymus DNA and poly(dG-dC) has been studied by 13C and 1H NMR with emphasis on the mode of binding, the role of Mg2+, and pH effects. The most prominent changes in the DNA base pair 13C NMR resonances upon complexation with chromomycin were observed for G and C bases, consistent with the G-C preference exhibited by this compound. Comparison of the 13C spectrum of DNA-bound chromomycin A3 with that of DNA-bound actinomycin D, a known intercalator, showed many similarities in the base pair resonances. This suggested the possibility that chromomycin A3 binds via an intercalative mechanism. 1H NMR studies in the imino proton, low-field region of the spectrum provided additional evidence in support of this binding mode. In the low-field spectrum of chromomycin A3 bound to calf thymus DNA, a small shoulder was observed on the upfield side of the G-C imino proton peak. Similarly, in the chromomycin A3 complex with poly(dG-dC), a well-resolved peak was found upfield from the G-C imino proton peak. These results are expected for ligands that bind by intercalation. Furthermore, in both the calf thymus and poly(dG-dC) drug complexes (in the presence of Mg2+) a broad peak was also present downfield (approximately 16 ppm from TSP) from the DNA imino protons. This was attributed to the C-9 phenolic hydroxyl proton on the chromomycin chromophore. Visible absorbance spectra at different pH values showed that the role of Mg2+ in the binding of chromomycin A3 to DNA is more than simple neutralization of the drug's anionic change.

Animals↗

PubMed1990

Chromomycin dimer-DNA oligomer complexes. Sequence selectivity and divalent cation specificity.

This paper reports on a solution NMR characterization of the sequence selectivity and metal ion specificity in chromomycin-DNA oligomer complexes in the presence of divalent cations. The sequence selectivity studies have focused on chromomycin complexes with the self-complementary d(A1-A2-G3-G4-C5-C6-T7-T8) duplex containing a pair of adjacent (G3-G4).(C5-C6) steps and the self-complementary d(A1-G2-G3-A4-T5-C6-C7-T8) duplex containing a pair of separated (G2-G3).(C6-C7) steps in aqueous solution. The antitumor agent (chromomycin) and nucleic acid protons have been assigned following analysis of distance connectivities in NOESY spectra and coupling connectivities in DQF-COSY spectra for both complexes in H2O and D2O solution. The observed intermolecular NOEs establish that chromomycin binds as a Mg(II)-coordinated dimer [1 Mg(II) per complex] and contacts the minor-groove edge with retention of 2-fold symmetry centered about the (G3-G4-C5-C6).(G3-G4-C5-C6) segment of the d(A2G2C2T2) duplex. By contrast, complex formation is centered about the (G2-G3-A4-T5).(A4-T5-C6-C7) segment and results in removal of the two fold symmetry of the d(AG2ATC2T) duplex. Thus, the binding of one subunit of the chromomycin dimer at its preferred (G-G).(C-C) site assists in the binding of the second subunit to the less preferred adjacent (A-T).(A-T) site. These observations suggest a hierarchy of chromomycin binding sites, with a strong site detected at the (G-G) step due to the hydrogen-bonding potential of acceptor N3 and donor NH2 groups of guanosine that line the minor groove. The divalent cation specificity has been investigated by studies on the symmetric chromomycin-d(A2G2C2T2) complex in the presence of diamagnetic Mg(II), Zn(II), and Cd(II) cations and paramagnetic Ni(II) and Co(II) cations. A comparative NOESY study of the Mg(II) and Ni(II) symmetric complexes suggests that a single tightly bound divalent cation aligns the two chromomycins in the dimer through coordination to the C1 carbonyl and C9 enolate ions on the hydrophilic edge of each aglycon ring. Secondary divalent cation binding sites involve coordination to the major-groove N7 atoms on adjacent guanosines in G-G steps. This coordination is perturbed on lowering the pH below 6.0, presumably due to protonation of the N7 atoms. The midpoint of the thermal dissociation of the symmetric complex is dependent on the divalent cation with the stability for reversible transitions decreasing in the order Mg(II) greater than Zn(II) greater than Cd(II) complexes.(ABSTRACT TRUNCATED AT 400 WORDS)

Base Sequence↗

PubMed1977

R-banding produced by DNase I digestion of chromomycin-stained chromosomes.

A distinct reverse (R-) banding pattern was produced on human chromosomes by digesting chromosome spreads with pancreatic deoxyribonuclease I (DNase I) in the presence of an excess of chromomycin A3 (CMA), followed by staining with Giemsa. The banding pattern corresponds with that obtained by chromomycin A3 fluorescence, and bands which fluorescence brightly with chromomycin appear darkly with Giemsa. The same relationship was observed in two plants, Scilla siberica and Ornithogalum caudatum, which have contrasting types of heterochromatin. Chromomycin bright C-bands stained darkly with the CMA/DNase I technique, whereas chromomycin negative C-bands appeared lightly stained. The digestion patterns are thought to reflect the variation in chromomycin binding capacity along the chromosome with R-bands and dark C-bands being sites which preferentially bind the antibiotic.

Azure Stains↗

PubMed1988

Solution conformation of the antitumor antibiotic chromomycin A3 determined by two-dimensional NMR spectroscopy.

A conformational analysis and a complete assignment of the nonexchangeable proton resonances of chromomycin A3, dechromose-A chromomycin A3, and deacetylchromose-B chromomycin A3 were carried out in organic solvents. The resulting conformation in methanol has the three side chains of chromomycin A3 fully extended, away from one another and from the aglycon. In dichloromethane on the other hand, the drug was shown to adopt a highly compact conformation in which most of the 26 oxygen atoms in the molecule point out toward the solvent. The two carbohydrate side chains extend parallel to each other on the same side of the aglycon. Two intramolecular nuclear Overhauser enhancement contacts have been observed between different sugar units on these side chains, indicating close proximity for these moieties. In addition, the aliphatic side chain is folded toward the aglycon, parallel to the two oligosaccharide side chains. The overall conformation has a wedge-like shape with the two phenoxy groups exposed at the pointed edge. The presence of some exchange cross-peaks in the NOESY spectra suggests the presence of intramolecular hydrogen bonds that probably help to maintain the compact conformation. The derivatives of chromomycin A3 have qualitatively similar conformations, though their respective conformations are not as compact as the parent drug. The significance of these results is discussed in terms of a model of chromomycin A3 binding to DNA in the major groove.

Chromomycin A3↗

PubMed1988

Chromomycin A3 binds to left-handed poly(dG-m5dC).

The interaction of chromomycin A3 (an antitumor antibiotic) with right-handed and left-handed polynucleotides has been studied by absorbance, fluorescence, circular dichroism, 31P-NMR and 1H-NMR techniques. Binding to either the B form of poly(dG-dC) or the Z form of poly(dG-m5dC) shifts the absorbance maximum to higher wavelength and enhances the fluorescence of the drug. Circular dichroic spectra of solutions containing various concentrations of chromomycin A3 and fixed concentrations of either B or Z polynucleotides show well defined isoelliptic points at similar wavelengths. At the isoelliptic point, the drug complex with B DNA exhibits positive ellipticity while with Z DNA it exhibits negative ellipticity. 31P-NMR spectra of the chromomycin A3 complex with the Z form of poly(dG-m5dC) demonstrate that the Z conformation is retained in the drug complex up to one molecule drug/four base pairs. At Mg2+ concentrations lower than that necessary to stabilize the left-handed conformation of poly(dG-m5dC) alone, 31P analysis shows that chromomycin A3 can bind simultaneously to both the B and Z conformations of poly(dG-m5dC), with no effect on the B-Z equilibrium. These data demonstrate that chromomycin A3 binds to left-handed poly(dG-m5dC) with retention of the left-handed conformation up to saturating drug concentrations.

Binding Sites↗

PubMed1992

Structure refinement of the chromomycin dimer-DNA oligomer complex in solution.

We have refined the initial docking model of the Mg(II)-co-ordinated chromomycin-d(A2G2C2T2) complex (2 drug equivalents per duplex) by a complete relaxation matrix analysis simulation of the two-dimensional nuclear Overhauser effect (NOESY) spectrum of the complex in 2H2O solution. This relaxation matrix refined structure of the complex exhibits the following characteristics. (1) We observe an unwound and elongated duplex that exhibits characteristics distinct from the A and B-DNA family of helices at the central (G-G-C-C).(G-G-C-C) chromomycin dimer binding and flanking sites. On the other hand sugar puckers, glycosidic torsion angles, displacement of the base-pairs from the helix axis and the minor groove width for this central tetranucleotide segment all fall within the A-family of helical parameters. (2) The chromomycin monomers are aligned in a head-to-tail orientation in the Mg(II)-co-ordinated dimer in the complex. The chromophores are aligned with a slight tilt relative to each other and make an angle of 75 degrees between their planes. The C-D-E trisaccharide segments from individual monomers adopt an extended conformation that projects in opposite directions in the dimer. The divalent metal cation is co-ordinated to the O(1) carbonyl and O(9) enolate atoms of the chromophores and aligns them such that the O(9)-Mg-O(9) angle is 170 degrees while all other O-Mg-O angles are in the 95(+/- 15)degrees range. (3) The sequence specificity of the chromomycin dimer for the widened and shallower (G3-G4-C5-C6).(G3-G4-C5-C6) minor groove binding site is associated with intermolecular hydrogen bonds formed between the OH group at C(8) of the chromophore and the minor groove NH2 group at position 2 and N(3) groups of G4 and between the O(1) oxygen of the E-sugar and the minor groove NH2 group at position 2 of G3 in the complex. (4) Additional intermolecular interactions are primarily van der Waals contacts between anomeric and adjacent CH2 protons on each sugar in the C-D-E trisaccharide segments of the chromomycin dimer and the minor groove surface of the DNA. These results provide insights into the induced conformational transitions required to generate a complementary match between the drug dimer and its DNA binding site on complex formation.

Base Sequence↗

PubMed1977

Chromomycin A3 as a fluorescent probe for flow cytometry of human gynecologic samples.

Chemical, physical and optical properties of chromomycin A3 are examined so as to ascertain appropriate staining and analysis procedures for flow cytometry of human gynecologic samples. Fluorescence excitation and emission spectra of chromomycin A3-stained cervical cells are compared with those of chromomycin A3-stained deoxyribonucleic acid. Conditions for deoxyribonucleic acid-specific staining of cervical cells are presented, and staining specificity of cervical cells with chromomycin A3 is compared to that obtained with ethidium bromide, propidium iodide and Hoechst 33258. Also presented is a brief review of two parameter flow cytometry as a prescreening procedure for detection of cervical neoplasia. Results of flow cytometry and cell sorting are interpreted based on the deoxyribonucleic acid-specificity of chromomycin A3 staining.

Cell Nucleus↗

PubMed1989

Solution structure of the chromomycin-DNA complex.

The structure of the chromomycin-DNA complex at the deoxyoctanucleotide duplex level has been determined from one- and two-dimensional proton NMR studies in Mg-containing aqueous solution. The NMR results demonstrate that the antitumor agent binds as a symmetrical dimer to the self-complementary d[T-T-G-G-C-C-A-A] duplex with retention of the 2-fold symmetry in the complex. A set of intermolecular nuclear Overhauser enhancements (NOEs) establishes that two chromomycin molecules in the dimer share the minor groove at the G-G-C-C.G-G-C-C segment in such a way that each hydrophilic edge of the chromophore is located next to the G-G.C-C half-site and each C-D-E trisaccharide chain extends toward the 3'-direction of the octanucleotide duplex. In addition, the A-B disaccharide segment and the hydrophilic side chain of the antitumor agent are directed toward the phosphate backbone. The observed changes in nucleic acid NOEs and coupling patterns on complex formation establish a transition to a wider and shallower minor groove at the central G-G-C-C.G-G-C-C segment required for accommodating the chromomycin dimer. The present demonstration that chromomycin binds as a dimer and switches the conformation of the DNA at its G.C-rich minor groove binding site provides new insights into antitumor agent design and the sequence specificity of antitumor agent-DNA recognition.

Base Sequence↗

PubMed1990

NMR investigation of mithramycin A binding to d(ATGCAT)2: a comparative study with chromomycin A3.

The binding of mithramycin A to the d(A1T2G3C4A5T6) duplex was investigated by 1H NMR and found to be similar to that of its analogue chromomycin A3. In the presence of Mg2+, mithramycin binds strongly to d(ATGCAT)2. On the basis of the two-dimensional NOESY spectrum, the complex formed possesses C2 symmetry at a stoichiometry of two drugs per duplex (2:1) and is in slow chemical exchange on the NMR time scale. NOESY experiments reveal contacts from the E-pyranose of mithramycin to the terminal and nonterminal adenine H2 proton of DNA and from the drug hydroxyl proton to both G3NH2 protons, C4H1' proton, and A5H1' proton. These data place the drug chromophore and E pyranose on the minor groove side of d(ATGCAT)2. NOE contacts from the A-, B-, C-, and D-pyranoses of mithramycin to several deoxyribose protons suggest that the A- and B-rings are oriented along the sugar-phosphate backbone of G3-C4, while the C- and D-rings are located along the sugar-phosphate backbone of A5-T6. These drug-DNA contacts are very similar to those found for chromomycin binding to d(ATGCAT)2. Unlike chromomycin, the NOESY spectrum of mithramycin at the molar ratio of one drug per duplex reveals several chemical exchange cross-peaks corresponding to the drug-free and drug-bound proton resonances. From the intensity of these cross-peaks and the corresponding diagonal peaks, the off-rate constant was estimated to be 0.4 s-1. These data suggest that the exchange rate of mithramycin binding to d(ATGCAT)2 is faster than that of chromomycin.

Base Sequence↗

PubMed1975

[Polymorphism of a culture of the chromomycin producer, Actinomyces aburaviensis var. verrucosus].

Polymorphism of the chromomycin-producing orgnaism Act. aburaviensis var. verrucosus 144--3 was studied. The stable variants differed in the morphologo-cultural properties, assimilation of the carbon sources, the component composition of the luminescence substances and the quantitative property of the antibiotic production. A substance with violet luminescence was found in the culture of varient I in addition to chromomycin. Similarity of the phenotypes observed in variants 1 and 2 when grown on complex organic media is explained by intensive chromomycin production by variant 1 on such media. Active colonies may be selected according to the colour due to chromomycin. Forms with an activity almost 4 times higher than that of the initial culture were found among the colonies of variant 2.

Chromomycins↗

PubMed1991

An NMR study on the conformation of a deoxyoligonucleotide duplex, d(GGGGCCCC)2, and its complex with chromomycin.

The conformation of drug-free d(GGGGCCCC)2 and the chromomycin-d(GGGGCCCC)2 complex in aqueous solution were studied by NMR spectroscopy. The present study has indicated that free d(GGGGCCCC)2 takes the B form in solution, although it takes the A form in the crystalline state. The NMR spectrum of the complex indicated that chromomycin binds as a symmetry-related dimer to the minor groove of the central four residues of d(GGGGCCCC)2. The drastic conformational change in the central four residues of d(GGGGCCCC)2 on going from the B form family to the A form was demonstrated by the characteristic NOEs and coupling patterns. The change seems to be indispensable for accommodation of the bulky chromomycin dimer in the minor groove. On the basis of the intermolecular NOEs between chromomycin and d(GGGGCCCC)2, the structure of the complex has been constructed and refined by energy minimization.

Base Sequence↗

PubMed1990

An NMR study on the conformation of the chromomycin-d(GGGGCCCC)2 complex.

The conformation of the chromomycin-d(GGGGCCCC)2 complex in aqueous solution was studied by NMR spectroscopy. The NMR spectrum of the complex indicated that the chromomycin binds as a symmetry-related dimer to the minor groove of the central four residues of d(GGGGCCCC)2. The drastic conformational change in the central four residues of d(GGGGCCCC)2 from the B form family to the A-form was demonstrated by the characteristic NOEs and coupling patterns. The change seems to be indispensable for accommodation of the bulky chromomycin dimer in the minor groove. On the basis of the intermolecular NOEs between chromomycin and d(GGGGCCCC)2, the structure of the complex has been constructed and refined by energy minimization.

Chromomycins↗

PubMed1977

Combination chemotherapy for solid tumors using 5-fluorouracil, chromomycin-A, and prednisolone.

Clinical effect of 5-fluorouracil or chromomycin-A3 alone, 5-fluorouracil + chromomycin-A3, and of the first two plus prednisolone on gastrointestinal and other solid tumors was evaluated. Out of 133 cases acceptable for evaluation, the number of responders was as follows: 3 (18.8%) of 6 cases treated with 5-fluorouracil alone, 1 (9.1%) of 11 cases treated with chromomycin-A3 or chromomycin-A3 hemisuccinate, 13 (21.7%) of 60 cases on the two-drug regimen, and 21 (45.7%) of 46 cases on the three-drug regimen. In cases of stomach carcinoma, response rate to the three-drug regimen was 54.2% (13/24), significantly higher than that of other regimens. At least 25% regression in the size of primary tumor was observed in 2 (7.1%) of 28 cases on the two-drug regimen and in 6 (33.3%) of 18 cases on the three-drug regimen. Of 51 cases on the three-drug regimen, steroid diabetes developed in 5 cases, moon face in 4 cases, and gastric ulcer in 1 case. However, toxic effect of these regimens (especially appearance of leucopenia) was less than those of previously tried combined regimens. The duration of response, on an average, was 10.8 weeks in 13 cases on the two-drug regimen and 11.7 weeks in 21 cases on the three-drug regimen. It was concluded from these results that a better response is obtained by the three-drug regimen than other regimens, and that prednisolone in combination, in addition to its favorable effect in improving the general condition of the patients, might enhance the anticancer effect of the drugs used in combination.

Adenocarcinoma↗

PubMed1991

Proton exchange and internal motions in two chromomycin dimer-DNA oligomer complexes.

Previous structural studies on the complexes of the chromomycin (CHR) dimer with duplexes of d(A1-A2-G3-G4-C5-C6-T7-T8) and of d(A1-G2-G3-A4-T5-C6-C7-T8) in solution [one Mg(II) and two drugs per duplex] are extended to hydrogen exchange measurements. Exchange of the OH8 proton of chromomycin, measured by real time proton-deuterium exchange, is very slow and requires dissociation of the complex, whose lifetime is thus determined. The lifetimes and apparent dissociation constants of base pairs are deduced from the catalysis of imino proton exchange by ammonia. The four central base pairs, which interact with the CHR chromophores in the minor groove (Gao & Patel, 1990), may open within the complex, but the opening rate is less than in the free duplex by one to two orders of magnitude. The activation energy for base-pair opening and the differences between the lifetimes of adjacent pairs suggest that single base-pair opening is the predominant imino proton exchange pathway in all cases. In the symmetrical complex of chromomycin with the first duplex, the lifetimes of the central base pairs (G3.C6 and G4.C5) are in the same range (52 and 29 ms, respectively, at 38 degrees C). In the asymmetrical complex formed with the second duplex, the base-pair lifetimes in the G2-G3-A4-T5 segment that interacts with the chromophore moiety are strongly increased. That of G3.C6 is particularly long. Above 50 degrees C, exchange of the G3 imino proton is opening limited.(ABSTRACT TRUNCATED AT 250 WORDS)

Ammonia↗

PubMed1990

NMR studies of the interaction of chromomycin A3 with small DNA duplexes. Binding to GC-containing sequences.

The interaction of chromomycin A3 with the oligodeoxyribonucleotides 1, d(ATGCAT), 2, d(ATCGAT), 3, d(TATGCATA), and 4, d(ATAGCTAT), has been investigated by 1H and 31P NMR. In the presence of Mg2+, chromomycin binds strongly to the three GC-containing oligomers 1, 3, and 4 but not to the CG-containing oligomer 2. The proton chemical shift changes for 1 and 3 are similar, and these DNA duplexes appear to bind with a stoichiometry of 2 drugs:1 Mg2+:1 duplex. The same stoichiometry of 2 drugs:1 duplex is confirmed with 4; however, proton chemical shift changes differ. An overall C2 symmetry is exhibited by the drug complex with 1, 3, and 4. At a molar ratio of 2.0 (drugs:duplex), no free DNA proton NMR signals remain. Two-dimensional nuclear Overhauser exchange spectroscopy (NOESY) of the saturated chromomycin complex with 1 and 3 positions both chromomycinone hydroxyls and the E carbohydrates in the minor groove and provides evidence suggesting that the B carbohydrates lie on the major-groove side. This is supported by several dipolar coupling cross-peaks between the drug and the DNA duplex. Drug-induced conformational changes in duplex 1 are evaluated over a range of NOESY mixing times and found to possess some characteristics of both B-DNA and A-DNA, where the minor groove is wider and shallower. A widening of the minor groove is essential for the DNA duplex to accommodate two drug molecules. This current minor-groove model is a substantial revision of our earlier major-groove model [Keniry, M.A., Brown, S.C., Berman, E., & Shafer, R.H. (1987) Biochemistry 26, 1058-1067] and is in agreement with the model recently proposed by Gao and Patel [Gao, X., & Patel, D. J. (1989a) Biochemistry 28, 751-762].

Base Composition↗

PubMed1975

Mode of action of antitumour antibiotics. spectrophotometric studies on the interaction of chromomycin A3 with DNA and chromatin of normal and neoplastic tissue.

The binding of chromomycin A3, an antitumour antibiotic, to various DNA and chromatin isolated from mouse and rat liver, mouse fibrosarcoma and Yoshida ascites sarcoma cells was studied spectrophotometrically at 29 degrees C in 10-2 M Tris-HCl buffer, pH 8.0, containing small amounts of MgCl2 (4.5-10-5--25-10-5 M). An isobestic point at 415 nm was observed when chromomycin A3 was gradually titrated with DNA/chromatin and its spectrum shifted towards higher wavelength. The rates and extent of these spectral changes were found to be dependent on the concentration of Mg2+. The change in absorbance at 440 nm was used to calculate apparent binding constant (Kap M-1) and sites per nucleotide (n) from Scatchard plots for various DNA and chromatins. As expected, values of n for chromatin (0.06-0.10) were found to be lower than found for corresponding DNA (0.10-0.15). Apparently no such correlation exists between binding constants (Kap M-1)-10-4) of DNA (6.4--11.2) and of chromatin (3.1--8.3), but Kap M-1 of chromatin isolated from mouse fibrosarcoma and Yoshida ascites sarcoma are 1.5--3 times higher than that found for mouse and rat liver chromatin. These differences may be taken to indicate structural difference in nucleoprotein complexes caused by neoplasia. The relevance of this finding to tumour suppressive action of chromomycin A3 is discussed.

Animals↗