DNA-4'-6-diamidine-2-phenylindole interactions: a comparative study employing fluorescence and ultraviolet spectroscopy.

DAPI is a drug that interacts with double-stranded nucleic acids, binding preferentially to A + T base pairs. The interaction is not intercalative, therefore providing a useful model for mimicking the effect of functional molecules in modifying specific sites, namely, A + T segments, of significance in gene expression. Knowledge of the nature of such interaction has been enriched by additional information obtained from comparative analysis of the data acquired by uv spectroscopy and fluorescence. Two classes of binding sites, defined by different apparent affinity constants and numbers of binding sites, are evident. All types of interaction are dependent on the nucleic acid/dye ratio and on the ionic strength of the medium.

On the complex nature of the antiviral activity of coumermycin A1: its interference with the replication of herpes simplex virus type 1.

The mechanism of inhibition of the replication of herpes simplex virus type 1 (HSV-1) by coumermycin A1 (CA1), an inhibitor of bacterial DNA gyrase, has been investigated. Concentrations of antibiotic slightly higher than those needed for 50% inhibition of viral growth were able to inhibit viral DNA synthesis in infected cells. This effect was accompanied by a depressed synthesis of viral polypeptides. Protein synthesis was also inhibited in uninfected cells, especially after long exposure to the drug, but not in a cell-free system. In vitro assays of highly purified HSV-1 DNA polymerase in the presence of the drug, provided evidence that the enzyme was a target of CA1. The viral polymerase was in fact inhibited by the antibiotic to an extent comparable to that of viral DNA synthesis in intact cells. In contrast, DNA polymerase alpha, the enzyme involved in chromosomal DNA replication, was relatively insensitive to CA1. The drug was also shown to bind to protein and to viral and cellular DNA.

Interaction of papaverine with covalently closed DNA.

Cyclic nucleotides influence viral replication and papaverine, as an inhibitor of phosphodiesterase, also affects the replication of both DNA and RNA viruses through an increase in cAMP levels. Moreover in vitro papaverine affects neither protein synthesis nor several polymerases, while it inhibits DNA synthesis. Static fluorescence studies on the interaction of the drug with ColE1 plasmid covalently closed DNA indicate that the drug binds to the nucleic acid, probably by intercalation. A comparison between the binding characteristics of Papaverine and Actinomycin D is also reported.

Different ability of novobiocin and coumermycin A1 to interact with nucleic acids.

The possibility of two structurally related antibiotics, Coumermycin A1 and Novobiocin, to interact with nucleic acids was investigated. Only Coumermycin A1 was able to form complexes with DNA showing an apparent affinity constant comparable to that of the interaction with ribosomal RNA. A binding specificity for A + T complementary and repeating sequences was also exhibited by Coumermycin A1. In view of the different behaviour of the two compounds some considerations are made on their mode of action; although they are acting on the same target enzymes in Escherichia coli, they may affect the functions of eukaryotic cells through a different mechanism not equally specific and probably distinct for each of the two antibiotics.

Nature of toxicity for chick embryo fibroblast cells of coumermycin A1 and its physico-chemical interactions with protein and nucleic acid.

The results reported in this paper describe the effects produced by the antibiotic Coumermycin A1 (CA1) on survival and metabolism of chick embryo fibroblast cells (CEF), and give a clue to the understanding of its toxicity. The drug acts primarily at the level of DNA and RNA synthetic enzymes; no effect on DNA superstructure is detectable at doses at which cytotoxicity is pronounced. A spectroscopic approach produced evidence that CA1 binds to DNA, RNA, chromatin components such as histones and to a structurally unrelated protein such as bovine serum albumin. Furthermore, CA1 behaves like a pure non-competitive inhibitor of lactic dehydrogenase, a ubiquitous enzyme not involved in nucleic acid metabolism. The interaction of CA1 with a wide range of macromolecules playing different biological roles is certainly relevant to its activity and adds a new insight into the mechanism of action of this antibiotic. These observations are also discussed in the light of the alleged role of CA1 as a specific inhibitor of DNA topoisomerase in eukaryotic cells.

[Intermolecular interactions between fluorescent probes bound to linear DNA]. / Interazioni intermolecolari fra sonde fluorescenti legate a DNA lineare.

Intermolecular interactions of fluorescent probes bound to linear DNA. Several fluorescent probes can be widely used in determining the base content, the geometry and the possible conformational transitions of linear and circular DNAs in solution. Data are presented showing that Tb3+ and DAPI bind independently to DNA and can therefore be used to monitor the G+C and A+T content of the nucleic acid respectively.

Terbium as a new probe to detect G+C content of DNA.

Tb3+ has been tested as a probe to determine G + C per cent composition of DNA. DAPI, a fluorescent dye specific for A + T bases, has been employed at the same time as a complementary probe. Using calf thymus DNA (G + C = 40%; A + T = 60%), it has been found that the binding of the two dyes is independent, and that one binding site every 12,5 nucleotides exists for Tb3+ and one binding site every 8.3 nucleotides for DAPI. The simultaneous use of Tb3+ and DAPI allows therefore the monitoring of A-T, G-C sequences.

[Experimental study on hepatic encephalopathy: EEG and blood amino acid findings]. / "Studio sperimentale sull'encefalopatia epatica: rilievi EEGrafici ed aminoacidemici".

The AA. describe a method of gradual occlusion of the portal vein in the rat to induce a hepatic encephalopaty. This method allows to realise a condition of hepatic ischemia and a portal hypertensive state accompanied by spontaneous portal-systemic shunts. These two factors produce a hepatic encephalopaty like in cirrhotic state or in patients after portal systemic anastomosis. A part from similar behavior at the beginning of the encephalopaty it is possible to define two classes of animals: one showing a slow recovery and one showing a transient and slight improvement followed at the end by the death of the animal. The AA. found a definite correlation amoung the clinica, EEG's, serum aminoacids and biochemical data. Probably the different anatomical and phisiological aspects of the induced portal systemic shunts and the different ways of hepatic rivascularization may determine the two different evolutions of the animals.