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Biomedical subjects

A R Morgan

Publications and source records attributed to A R Morgan.

At least 73 records · Page 4Linked to original sources
PubMed1987

Morphological study of the combined effect of purpurin derivatives and light on transplantable rat bladder tumors.

Purpurin derivatives, a group of synthetic photosensitizers, were tested for their photodynamic activity against transplantable N-[4-(5-nitro-2-furyl)-2-thiazolyl]formamide-induced urothelial tumors growing in male Fischer 344 rats. Histological examination of tumors in animals treated with the purpurin derivatives and red light (greater than 590 nm, 360 joules/cm2) revealed tumor cell necrosis 24 h after completion of therapy. Control tumors showed no histological change. Purpurins have a major absorption peak in the red region (greater than 650 nm) of the visible spectrum, a region with good tissue penetration, and purpurins can be synthesized with a high degree of purity. This study demonstrates the potential of purpurin derivatives as photosensitizers for photodynamic cancer therapy.

Animals↗
PubMed1987

A dose response analysis of purpurin derivatives used as photosensitizers for the photodynamic treatment of transplantable FANFT induced urothelial tumors.

Rats engrafted with transplantable N-[4-(5-nitro-2-furyl)-2-thiazolyl] formamide (FANFT) induced urothelial tumors were treated with purpurin derivatives and red light (greater than 590 nm., 360 joules/cm.2). Treated and control tumors were harvested 12 days after treatment and dry weights compared. At doses of 5.0 micrograms./gm. and 2.5 micrograms./gm. body weight, the purpurins designated NT1 and NT2, when combined with light, caused statistically significant (p less than 0.02) tumor regression when compared to light shielded controls. At 1.0 micrograms./gm. body weight, NT1 and light also induced significant tumor regression (p less than 0.02). Purpurins, which have strong absorption bands above 650 nm. and can be synthesized with a high degree of purity, appear to have potential as photosensitizers for photodynamic therapy.

Animals↗
PubMed1986

Characterization of imperfect DNA duplexes containing unpaired bases and non-Watson-Crick base pairs.

Synthetic duplex DNAs of repeating sequence, such as poly d(TTC).poly d(GAA), were separated into their individual single strands. The various single strands complexed not only, as expected, with their complementary strands, but also with other non-complementary strands. Characterization of such complexes with respect to stoichiometry, Tm values and the dependence of Tm on NaCl concentration showed that a variety of unusual structures could be inferred at physiological salt concentrations. These included extrahelical thymines, G.T oppositions, A.C oppositions and T.C oppositions.

Base Composition↗
PubMed1986

The binding of the anthelmintic pyrvinium cation to deoxyribonucleic acid in vitro.

Pyrvinium pamoate, an anthelmintic drug with mutagenic activity, binds to duplex DNA in vitro. Binding markedly enhances the intrinsic fluorescence of the drug, permitting a characterization of the binding reaction to be performed directly through fluorimetric analysis. The monopyrvinium salt, pyrvinium iodide, binds cooperatively to native calf thymus DNA with an intrinsic binding constant of 1.1 X 10(4) M-1 as determined fluorimetrically and 1.4 X 10(4) M-1 as determined by equilibrium dialysis. The respective binding site was estimated to be 2.5 base pairs. A change in the absorption of bound drug as a function of the binding ratio identifies secondary, nonfluorescent binding which is essentially ionic in character. The binding of pyrvinium removes negative supercoils from PM2-ccc-DNA with an experimental efficiency greater than that observed for the intercalating drug chloroquine. Electron microscopy shows that molecules of pBR322 DNA treated with pyrvinium increase in length by as much as 18% over controls, consistent with an increase in DNA base pair separation upon binding. The evidence indicates that the primary interaction of pyrvinium with DNA involves intercalation.

Animals↗
PubMed1985

Characteristics of the binding of the anticancer agents mitoxantrone and ametantrone and related structures to deoxyribonucleic acids.

The binding constants for interaction of the anticancer agents mitoxantrone and ametantrone and several congeners with calf thymus DNA and the effects of ionic strength changes have been determined spectrophotometrically. The agents show a preference for certain sequences, particularly those with GC base pairs, and the magnitude of the specificity depends on the specific substituents on the anthraquinone ring system. The binding constant for mitoxantrone with calf thymus DNA in 0.1 M Na+, pH 7, is approximately 6 X 10(6) M-1, and the rate constant for the sodium dodecyl sulfate driven dissociation of mitoxantrone from its calf thymus DNA complex under the same solution conditions and 20 degrees C was determined to be 1.3 s-1. The unwinding angle of mitoxantrone determined independently by viscosity measurements and by a novel assay employing calf thymus topoisomerase shows excellent agreement for a value of 17.5 degrees. The viscosity increase of sonicated calf thymus DNA varies considerably with the substituent on the anthraquinone ring system. Binding studies employing T4 and phi w-14 DNAs in which the major groove is occluded and the reverse experiment with anthramycin-treated calf thymus DNA indicate at least part of the mitoxantrone molecule may lie in the minor groove.

Animals↗
PubMed1985

Restriction nuclease digestions driven to completion by Escherichia coli RNA polymerase and T4 gene 32 protein.

Restriction enzyme digestions of large scale DNA preparations often do not go to completion. This is due to product inhibition by the newly generated ends of the digested DNA. The addition of exogenous proteins that bind tightly to the free ends of DNA or to single-stranded DNA will relieve this inhibition. We show that a considerable savings on restriction nucleases can be attained by the addition of RNA polymerase or T4 gene 32 protein in stoichiometric amounts to the newly produced DNA ends.

DNA Restriction Enzymes↗
PubMed1985

A structural basis for S1 nuclease sensitivity of double-stranded DNA.

A protonated form of a cloned simple repeating DNA sequence d(TC)n X d(GA) is detectable in equilibrium with the usual Watson-Crick base-paired form at pHs up to 7. This form is anomalously sensitive to a variety of single-strand-specific endonucleases. The observed pH dependent protection of N-7 of dG residues within the insert suggests that these residues are either Hoogsteen or reverse Hoogsteen base-paired to protonated dC residues of the polypyrimidine strand. A structure in which dA:dT Watson-Crick base pairs alternate with Hoogsteen syndG:dCH+ pairs appears to be the most stereochemically acceptable structure consistent with the chemical properties of this protonated DNA. Protonated d(TC)n X d(GA)n interacts with an anti-Z DNA antibody raised against brominated d(GC)n X d(GC)n.

Cloning, Molecular↗
PubMed1984

Poly(pyrimidine) . poly(purine) synthetic DNAs containing 5-methylcytosine form stable triplexes at neutral pH.

Poly(pyrimidine) . poly(purine) tracts have been discovered in the 5'-flanking regions of many eucaryotic genes. They may be involved in the regulation of expression since they can be mapped to the nuclease-sensitive sites of active chromatin. We have found that poly(pyrimidine) . poly(purine) DNAs which contain 5-methylcytosine (e.g. poly[d(Tm5C)] . poly[d(GA)]) will form a triplex at a pH below 8. In contrast, the unmethylated analogue, poly[d(TC)] . poly[d(GA)] only forms a triplex at pHs below 6. Synthetic DNAs containing repeating trinucleotides and poly[d(Um5C)] . poly[d(GA)] behave in a similar manner. Thus the stability of a triplex can be controlled by methylation of cytosine. This suggests a model for the regulation of expression based upon specific triplex formation on the 5'-side of eucaryotic genes.

5-Methylcytosine↗
PubMed1984

Rapid sensitive fluorescence assays for DNA endonucleases and DNA glycosylases.

Using ethidium fluorescence as a probe and the unique topological properties of covalently closed circular (CCC) DNA's picogram quantities of pancreatic DNase I can be detected. By chemically modifying CCC DNA's without the introduction of nicks, various DNA repair enzymes can be detected even in crude cell extracts. This has previously been done for AP endonucleases from yeast. In this paper we describe the formation of ultraviolet (UV) and bisulfite modified CCC DNA's for the detection of T4 UV endonuclease (a DNA glycosylase + AP endonuclease) and uracil-DNA glycosylases, respectively. These assays should be useful in the isolation of DNA repair mutants.

DNA Glycosylases↗
PubMed1983

Studies of the interaction of RecA protein with DNA.

Ethidium fluorescence assays were adapted for the rapid and sensitive detection of precA; in addition, fluorescence measurements on binding precA to linear, OC and CCC PM2 DNAs have enabled the stoichiometry of precA binding as well as the precA-induced unwinding angle of DNA to be determined. The stoichiometry of binding was independently confirmed by sedimentation analysis to be one precA molecule per 3 bp. The unwinding angle was also independently confirmed by measurements of fluorescence changes induced by the binding of precA to CCC DNA which was relaxed by topoisomerase to give a precA-induced unwinding angle of 51 degrees. Electron microscopy of OC DNA molecules which bound nonsaturating amounts of precA revealed that the length increase in DNA due to precA was approximately 55%. Finally, examination of negatively stained precA complexes with a variety of linear DNAs showed that the minor groove is the primary site of interaction for this protein.

DNA Topoisomerases, Type I↗
PubMed1983

Left-handed Z-DNA regions are present in negatively supercoiled bacteriophage PM2 DNA.

Bacteriophage PM2 DNA is a 10-kb covalently closed circular (ccc) molecule with a reported superhelical density of sigma = -0.12. Here we describe the binding of anti-Z-DNA antibodies to PM2 form I DNA under high and low salt conditions. The binding to PM2 DNA has been demonstrated by competitive radioimmunoassay (RIA), retardation of the DNA:antibody complexes in agarose gels and visualization by electron microscopy. The antibody binding is dependent on the degree of negative supercoiling. Thus, PM2 form II and form III did not bind the antibody. The low salt RIA results indicated the presence of 200-400 bp of left-handed DNA per PM2 molecule. This could reduce the effective superhelical density to sigma = -0.04 to -0.08, a range comparable with those found for other ccc DNAs in vivo. Electron microscopy revealed that a maximum of 22 antibody molecules bind to PM2. Single-site restriction with HpaII of the fixed DNA:antibody complex showed a cluster of four to five antibody molecules bound near one end of the linear DNA molecule. The evidence presented indicates that PM2 DNA contains regions of left-handed conformation under physiological conditions (low salt concentration) as well as at high salt concentrations. In addition, electrophoretic analyses of PM2 topoisomers indicate the presence of left-handed regions at superhelical densities less than that of isolated PM2 DNA.

Antigen-Antibody Complex↗
PubMed1982

A method for the specific inhibition of poly[d(A-T)] synthesis using the A-T specific quinoxaline antibiotic TANDEM.

A serious problem in the replication of repeating-sequence DNa polymers using Escherichia coli DNA polymerase I arises from the fact that this polymerase has a very strong preference for the replication of poly[d(A-T)]. Thus reactions primed with DNA containing small amounts of contaminating poly[d(A-T)] will eventually result in complete domination of the synthesis by poly[d(A-T)]. This problem can be overcome by the addition to the reaction mixture of the synthetic quinoxaline antibiotic TANDEM which binds specifically to poly[d(A-T)] completely inhibiting its replication. Using thermal denaturation experiments it can be shown that TANDEM does not bind to most other synthetic DNA polymers (e.g., poly(dA) . poly(dT) and poly[d(A-T-C)] . poly[d(G-A-T)] and therefore their replication is not inhibited. The only exception we have encountered is poly[d(T-A-C)] . poly[d(G-T-A)] which does bind TANDEM and therefore the drug cannot be used during the synthesis of this polymer. The fact that poly[d(T-A-C)] . poly[d(G-T-A)] does bind TANDEM while poly[d(A-T-C)] . poly[d(G-A-T)] does not, suggests that the drug recognizes T-A rather than A-T sequences.

Anti-Bacterial Agents↗
PubMed1982

Novel aspects of the structure of the Escherichia coli nucleoid investigated by a rapid sedimentation assay.

The Escherichia coli nucleoid is maintained in its folded highly condensed state by constraints which involve RNA and protein. We have developed a rapid sedimentation assay to determine the state of folding of the membrane-free nucleoid. An approximate measure of the stability of the nucleoids under various conditions can then be estimated by measuring the temperature at which the nucleoids unfold. Using ethidium and gamma irradiation (which removes the negative supercoiling of the native nucleoid) as probes, it can be shown that there are two types of constraint involved in the condensation of the nucleoid. One of these constraints is destabilized by ethidium but stabilized by negative supercoiling; the second constraint is unaffected by both ethidium and negative supercoiling. Several models can be proposed: (i) a DNA . RNA duplex, (ii) a double-strand DNA (dsDNA) . RNA triplex, (iii) DNA-protein interactions, (iv) a topological knot with RNA, and (v) a DNA tetraplex. The topological knot model is not consistent with the data and many combinations of the others can be excluded. If RNA is involved in both constraints then RNA . DNA duplexes and dsDNA . RNA triplexes are involved in stabilizing the nucleoid.

Bacterial Proteins↗
PubMed1981

Monoclonal antibodies showing sequence specificity in their interaction with single-stranded DNAs.

Six hybridoma cell lines which secrete monoclonal antibodies binding to nucleic acids were produced from autoimmune NZB/NZW mice. Four of the antibodies were IgG's and the other two were IgM's. Using a solid phase radioimmunoassay (SPRIA) the binding of the antibodies to over thirty different nucleic acids was estimated. All the antibodies were extremely specific. There was no detectable interaction with various RNAs, and single-stranded DNAs bound more antibodies than duplex or multi-stranded DNAs. In every case the antibodies also showed considerable sequence preferences. For example one monoclonal antibody bound to d(TTC)n but not to d(TCC)n while another interacted strongly with D(TG)n and d(CA)n but not with d(TC)n, d(GA)n or homopolymers. In other cases the patterns of sequence specificity were extremely difficult to interpret although it seems clear that monoclonal antibodies have the potential to distinguish between any two nucleic acids however similar.

Animals↗
PubMed1980

Polypurine DNAs and RNAs form secondary structures which may be tetra-stranded.

Polypurine DNAs and RNAs containing at least 33% guanine form a stable secondary structure at neutral pH and moderate ionic strengths. The tm's of the polymers increase with increasing guanine content. To eliminate possible structures three novel polymers, d(Gn2A)n, d(Gm6A)n and d(IA)n, as well as the random copolymer rr(G,A)n were were studied. Both d(Gn2A)n and d(IA)n can form a secondary structure whereas d(Gm6A)n and r(G,A)n cannot. Model building suggested two possible structures, one a duplex and the other a tetra-stranded polymer. The latter is considered to be the more likely, since previous X-ray diffraction studies have shown that rGn and rIn are tetra-stranded. Circular dichroism spectra are also consistent with such an interpretation.

DNA↗