EVIDENCE FOR A NUCLEOTIDE-DEPENDENT TOPOISOMERASE ACTIVITY FROM YEASTMITOCHONDRIA

Yeast mitochondria were found to contain a novel topoisomerase-like ac tivity which required nucleoside di- or tri-phosphates as a cofactor. ADP supported activity as effectively as ATP and the optimal concentra tion for each was approximately 20 mu M. None of the other standard ri bo- or deoxyrib-onucleotides could fully substitute for either ADP or ATP. The non-hydrolyzable ATP analogs, adenosine-5'-O-(3-thiotriphosph ate) (ATP-gamma-S), adenylyl(beta,gamma-methylene) (AMP-PCP), and ande nyl-imidodiphosphate (AMP-PNP) also supported activity suggesting that the nucleotide cofactor regulated topoisomerase activity rather than serving as an energy donor in the reaction. The mitochondrial topoisom erase activity relaxed both positively and negatively supercoiled DNA. It was not inhibited by concentrations of ethidium bromide up to 2 mu g/ml nor by either nalidixic or oxolinic acids; novobiocin, coumermyc in, and berenil inhibited the activity. Genetic and biochemical analys is of the mitochondrial topoisomerase activity indicated that it was n ot encoded by the nuclear TOP1, TOP2, and TOP3 genes.