Relationship between DNA methylation and mutational patterns induced by a sequence selective minor groove methylating agent.
(1/132)
Me-lex, a methyl sulfonate ester appended to a neutral N-methylpyrrolecarboxamide-based dipeptide, was synthesized to preferentially generate N3-methyladenine (3-MeA) adducts which are expected to be cytotoxic rather than mutagenic DNA lesions. In the present study, the sequence specificity for DNA alkylation by Me-lex was determined in the p53 cDNA through the conversion of the adducted sites into single strand breaks and sequencing gel analysis. In order to establish the mutagenic and lethal properties of Me-lex lesions, a yeast expression vector harboring the human wild-type p53 cDNA was treated in vitro with Me-lex, and transfected into a yeast strain containing the ADE2 gene regulated by a p53-responsive promoter. The results showed that: 1) more than 99% of the lesions induced by Me-lex are 3-MeA; 2) the co-addition of distamycin quantitatively inhibited methylation at all minor groove sites; 3) Me-lex selectively methylated A's that are in, or immediately adjacent to, the lex equilibrium binding sites; 4) all but 6 of the 33 independent mutations were base pair substitutions, the majority of which (17/33; 52%) were AT-targeted; 5) AT --> TA transversions were the predominant mutations observed (13/33; 39%); 6) 13 out of 33 (39%) independent mutations involved a single lex-binding site encompassing positions A600-602 and 9 occurred at position 602 which is a real Me-lex mutation hotspot (n = 9, p < 10(-6), Poisson's normal distribution). A hypothetical model for the interpretation of mutational events at this site is proposed. The present work is the first report on mutational properties of Me-lex. Our results suggest that 3-MeA is not only a cytotoxic but also a premutagenic lesion which exerts this unexpected property in a strict sequence-dependent manner. (+info)
Specific molecular recognition of mixed nucleic acid sequences: an aromatic dication that binds in the DNA minor groove as a dimer.
(2/132)
Phenylamidine cationic groups linked by a furan ring (furamidine) and related compounds bind as monomers to AT sequences of DNA. An unsymmetric derivative (DB293) with one of the phenyl rings of furamidine replaced with a benzimidazole has been found by quantitative footprinting analyses to bind to GC-containing sites on DNA more strongly than to pure AT sequences. NMR structural analysis and surface plasmon resonance binding results clearly demonstrate that DB293 binds in the minor groove at specific GC-containing sequences of DNA in a highly cooperative manner as a stacked dimer. Neither the symmetric bisphenyl nor bisbenzimidazole analogs of DB293 bind significantly to the GC containing sequences. DB293 provides a paradigm for design of compounds for specific recognition of mixed DNA sequences and extends the boundaries for small molecule-DNA recognition. (+info)
Sequence-specific binding of counterions to B-DNA.
(3/132)
Recent studies by x-ray crystallography, NMR, and molecular simulations have suggested that monovalent counterions can penetrate deeply into the minor groove of B form DNA. Such groove-bound ions potentially could play an important role in AT-tract bending and groove narrowing, thereby modulating DNA function in vivo. To address this issue, we report here (23)Na magnetic relaxation dispersion measurements on oligonucleotides, including difference experiments with the groove-binding drug netropsin. The exquisite sensitivity of this method to ions in long-lived and intimate association with DNA allows us to detect sequence-specific sodium ion binding in the minor groove AT tract of three B-DNA dodecamers. The sodium ion occupancy is only a few percent, however, and therefore is not likely to contribute importantly to the ensemble of B-DNA structures. We also report results of ion competition experiments, indicating that potassium, rubidium, and cesium ions bind to the minor groove with similarly weak affinity as sodium ions, whereas ammonium ion binding is somewhat stronger. The present findings are discussed in the light of previous NMR and diffraction studies of sequence-specific counterion binding to DNA. (+info)
Synthesis and DNA-binding of acridine-netropsin hybrid molecules.
(4/132)
We have designed and synthesized acridine-netropsin hybrid molecules. Spectroscopic (absorption, CD, flow dichroism and fluorescence) measurements reveal that hybrid molecules interact with DNA by both intercalation and minor-groove binding and shows enhanced preference for AT-rich sites. (+info)
Conformation dependent binding of netropsin and distamycin to DNA and DNA model polymers.
(5/132)
The binding of the antibiotics netropsin and distamycin A to DNA has been studied by thermal melting, CD and sedimentation analysis. Netropsin binds strongly at antibiotic/nucleotide ratios up to at least 0.05. CD spectra obtained using DNA model polymers reveal that netropsin binds tightly to poly (dA) . poly (dT), poly (dA-dT) . poly(dA-dT) and poly (dI-dC) . poly (dI-dC) but poorly, if at all, to poly (dG) . poly (dC). Binding curves obtained with calf thymus DNA reveal one netropsin-binding site per 6.0 nucleotides (K(a)=2.9 . 10(5) M(-1)); corresponding values for distamycin A are one site per 6.1 nucleotides with K(a)= 11.6 . 10(5) M(-1). Binding sites apparently involve predominantly A.T-rich sequences whose specific conformation determines their high affinity for the two antibiotics. It is suggested that the binding is stabilized primarily by hydrogen bonding and electrostatic interactions probably in the narrow groove of the DNA helix, but without intercalation. Any local structural deformation of the helix does not involve unwinding greater than approximately 3 degrees per bound antibiotic molecule. (+info)
Potent inhibition of werner and bloom helicases by DNA minor groove binding drugs.
(6/132)
Maintenance of genomic integrity is vital to all organisms. A number of human genetic disorders, including Werner Syndrome, Bloom Syndrome and Rothmund-Thomson Syndrome, exhibit genomic instability with some phenotypic characteristics of premature aging and cancer predisposition. Presumably the aberrant cellular and clinical phenotypes in these disorders arise from defects in important DNA metabolic pathways such as replication, recombination or repair. These syndromes are all characterized by defects in a member of the RecQ family of DNA helicases. To obtain a better understanding of how these enzymes function in DNA metabolic pathways that directly influence chromosomal integrity, we have examined the effects of non-covalent DNA modifications on the catalytic activities of purified Werner (WRN) and Bloom (BLM) DNA helicases. A panel of DNA-binding ligands displaying unique properties for interacting with double helical DNA was tested for their effects on the unwinding activity of WRN and BLM helicases on a partial duplex DNA substrate. The levels of inhibition by a number of these compounds were distinct from previously reported values for viral, prokaryotic and eukaryotic helicases. The results demonstrate that BLM and WRN proteins exhibit similar sensitivity profiles to these DNA-binding ligands and are most potently inhibited by the structurally related minor groove binders distamycin A and netropsin (K(i) +info)
Molecular modelling, synthesis and antitumour activity of carbocyclic analogues of netropsin and distamycin--new carriers of alkylating elements.
(7/132)
A series of netropsin and distamycin analogues was synthesised and investigated by molecular modelling. The lowest-energy conformations of four carbocyclic lexitropsins, potential carriers of alkylating elements, were obtained using the HyperChem 4.0 program, and compared with the DNA-lexitropsin crystal structures from the Brookhaven National Laboratory Protein Data Bank. A method for synthesis of carbocyclic lexitropsins was elaborated, with the use of a nitro group or azobenzene as precursors for the aromatic amino group. The influence of methoxy group in ortho position with respect to amide groups on the activity of the new compounds was investigated. All of the compounds tested showed high antitumour activity in the standard cell line of mammalian tumour MCF-7. (+info)
Synthetic analogues of netropsin and distamycin. VI. Synthesis of carbocyclic lexitropsins containing a bioreductive element.
(8/132)
Carbocyclic derivatives of lexitropsines containing of two aromatic rings, N-dimethylaminopropyl group lonked to carboxyl terminus and 5-[bis(2-chloroethyl)-amino]-2,4-dinitrobenzamide group linked to the amino terminus group were synthesed. The N-terminal group should present selective alkylating activity on the DNA of cancer cells in conditions of hypoxia. (+info)