coumermycin and diminazene-aceturate

A mitochondrial DNA topoisomerase (type I, ATP-independent) can be biochemically distinguished from the nuclear enzyme DNA topoisomerase I. This conclusion is based on the subcellular localization of the mitochondrial enzyme, its optimal reaction conditions and sensitivity to enzyme inhibitors. Unlike its nuclear counterpart, the mitochondrial DNA topoisomerase exhibits an absolute requirement for a divalent cation (Mg2+ and Ca2+ work equally well in vitro). In addition, it is slightly more sensitive to monovalent salts, with optimal activity obtained in 50-100 mM KCl. The mitochondrial enzyme is equally active at pH 7.5 or pH 9.5, but unlike its nuclear equivalent, is inactivated at higher pH values. The mitochondrial DNA topoisomerase is sensitive to coumermycin, berenil, camptothecin and 2,2,5,5-tetramethyl-4-imidazolidinone, a chemical that has no inhibitory effect on DNA topoisomerase I. Immunoblotting studies show that mitochondrial DNA topoisomerase activity is associated with a polypeptide (M(r) approximately 79,000) that cross-reacts with the antiserum against DNA topoisomerase I. Thus, the mitochondrial DNA topoisomerase may be derived by the differential expression of the DNA topoisomerase I gene or from the expression of a gene that is homologous to the DNA topoisomerase I gene.

Topics: Aminocoumarins; Camptothecin; Coumarins; Diminazene; Disinfectants; DNA Topoisomerases, Type I; Imidazoles; Mitochondria; Saccharomyces cerevisiae; Topoisomerase I Inhibitors

A DNA topoisomerase has been purified from vaccinia virus cores. The native enzyme is composed of a single subunit of 32,000 daltons. The enzyme relaxes both positively and negatively supercoiled DNA in the absence of an energy cofactor. Enzymatic activity is stimulated by magnesium ions and inhibited by ATP, and during relaxation the topoisomerase changes the linking number of the DNA substrate by steps of one. Trapping of the covalent DNA-enzyme intermediate has been achieved, and analysis of the cleavage of duplex DNA by the enzyme reveals that it makes a single-strand break and forms a covalent bond through the 3'-end of the broken strand. Enzymatic activity and formation of the trapped intermediate are inhibited by the drugs novobiocin, coumermycin A1, and berenil. The virally encapsidated enzyme has novel properties but most closely resembles a eucaryotic type I topoisomerase.

Topics: Adenosine Triphosphate; Aminocoumarins; Cells; Coumarins; Diminazene; DNA Topoisomerases, Type I; DNA, Superhelical; Eukaryotic Cells; Magnesium; Molecular Weight; Nalidixic Acid; Novobiocin; Vaccinia virus