Two-dimensional NMR studies on the anthramycin-d(ATGCAT)2 adduct (original) (raw)
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Journal of Medicinal Chemistry, 1988
Tomaymycin is a member of the pyrrole [ 1,4]benzodiazepine antitumor-antibiotic group that binds covalently to the exocyclic 2-amino group of guanine in DNA. Previous correlation of fluorescence and NMR data suggested that the 11R,llaS and the 11S,llaS diastereomers of tomaymycin could bind to DNA in two orientations relative to the covalently modified guanine (Barkley, M. D.; Cheatham, S.; Thurston, D. E.; Hurley, L. H. Biochemistry 1986,25,3021-3031). We now report on fluorescence, one-and two-dimensional proton NMR, and molecular modeling studies of the tomaymycin-d(ATGCAT)2 adduct, which corroborate these earlier observations. Fluorescence measurements show that there are two species of tomaymycin bound to d(ATGCAT)2, which are tentatively identified as the 11R,llaS and 11S,llaS diastereomers. Two distinct sets of signals for the tomaymycin molecule are present in the proton NMR spectrum of the tomaymycin-d(ATGCAT)2 duplex adduct. Two-dimensional correlation spectroscopy (2D-COSY) studies also show connectivities for four cytosine H5-H6 and eight thymine methyl-H6 protons and thus clearly establish the presence of two distinct species of tomaymycin4(ATGCAT)2 adducts in solution. A single scalar 11-lla lH NMR coupling in the 2D-COSY spectrum is indicative of an adduct species that has an S configuration a t the C-11 position. Two-dimensional nuclear Overhauser effect (NOESY) spectra of the tomaymycin-d(ATGCAT)2 duplex adduct show that the adducts are relatively nondistortive. In a NOESY experiment, cross-peaks were identified between both the aromatic H9 proton and the ethylidine methyl protons of tomaymycin and two different adenine H2 protons of d(ATGCAT),. Molecular mechanics calculations with AMBER show that the two species with the thermodynamically most favorable binding energies are the 11R,llaS and 11S,llaS isomers with their aromatic rings to the 5' and 3' sides of the covalently bound guanine, respectively. The NOES observed between tomaymycin protons and adenine H2 protons are in accord with molecular modeling studies. Taken together, these results strongly suggest that the two forms of tomaymycin bound to d(ATGCAT)2 are the 11S,llaS and 11R,llaS species, oriented with their aromatic rings to the 3' and 5' sides, respectively, of the covalently modified guanines.
The reaction of anthramycin with DNA
Biochimica et Biophysica Acta (BBA) - Nucleic Acids and Protein Synthesis, 1974
Anthramycin is known to react firmly with DNA, but no chemical basis for this interaction has been elucidated. Since anthramycin readily undergoes hydrolytic changes at C-11, the possible relation of this reaction to the interaction with DNA was considered. Both the hydrolysis of an ll-methoxy group and the reaction with DNA were found to be H * catalysed, thus supporting a relation between the two processes. The titration of H รท from the phenolic group at Position 9 of anthramycin was found to be absent in the DNA complex, suggesting that Position 9 also is involved in the complex. The rate of reaction of anthramycin-ll-methyl ether with DNA was found to be increased by prior hydrolysis. The rate of reaction with DNA, however, does not involve a rate-limiting conversion of anthramycin to a reactive component, since the kinetics of the reaction were found to be basically bimolecular. This excluded the possibility that a slow conformational change of the DNA was rate-limiting. The findings support the proposal that the binding of anthramycin to DNA is covalent, and that Positions 11 and 9 are involved in the binding. The bimolecular rate constant was found to decrease exponentially with the extent of DNA reaction, probably due to neighboring-site exclusion effects. The site of reaction involves guanine, since only guanine-containing deoxypolynucleotides were found to be reactive, and since reaction of DNA with anthramycin caused stochiometric elimination of binding sites of actinomycin.
Biochemistry, 1992
The interaction of thioformyldistamycin, an amide isostere of the naturally occurring antibiotic distamycin A, with a self-complementary decadeoxynucleotide duplex, d(CGCAATTGCG)2, has been examined using a variety of high-field 'H-NMR techniques. The ligand exhibits two forms in solution arising from geometric isomerism due to restricted rotation around the thioformamide bond. Only the thermodynamically more stable Z-form is shown to bind to the oligonucleotide along its minor groove at the central 5'-AATT segment with the end groups of the ligand extending into the flanking G C regions but without any close contact at the amidinium terminus. Cross-peaks involving characteristic intra-and interresidue proton connectivities in the 2D experiments (COSY and NOESY) were employed to assign individual resonances of both strands in the asymmetric DNA-drug complex. The solution structure of the complex was constructed by molecular mechanics calculations based upon initial estimates of drug-DNA NOE contacts and further refined through energy minimization. These results complement previous structural studies on distamycin and other lexitropsins with oligonucleotides. The exchange of the ligand between two equivalent binding sites on the DNA sequence was estimated to occur at 40 s-' with a free energy of activation of 16.5 kcal-mol-' at 321-326 K. There was no evidence of formation of a 2:l drug-oligomer complex, in contrast to the case of the natural product, which is attributed to steric demands of the larger sulfur atom. s e v e r a l compounds including distamycin A (Arcamone et al., 1967; Hahn, 1975), netropsin (Julia & Preau-Joseph, 1963), anthelvencin A (Probst et al., 1965), noformycin (Diana, 1973), and the kikumycins A and B (Takahishi et al.,
Anthracycline binding to DNA. High-resolution structure of d(TGTACA) complexed with 4'-epiadriamycin
European Journal of Biochemistry, 1992
Crystallographic methods have been applied to determine the high-resolution structure of the complex formed between the self-complementary oligonucleotide d(TGTACA) and the anthracycline antibiotic 4-epiadriamycin. The complex crystallises in the tetragonal system, space group P4,212 with c1 = 2.802 nm and c = 5.293 nm, and an asymmetric unit consisting of a single D N A strand, one drug molecule and 34 solvent molecules. The refinement converged with an R factor of 0.17 for the 2381 reflections with F a 3oF in the resolution range 0.70-0.14 nm. Two asymmetric units associate such that a distorted B-DNA-type hexanucleotide duplex is formed incorporating two drug molecules that are intercalated at the TpG steps. The amino sugar of 4'-epiadriamycin binds in the minor groove of the duplex and displays different interactions from those observed in previously determined structures. Interactions between the hydrophilic groups of the amino sugar and the oligonucleotidc are all mediated by solvent molecules. Ultraviolet melting measurements and comparison with other anthracycline-DNA complexes suggest that these indirect interactions have a powerful stabilising effect on the complex.