USE OF HAEMOPHILUS-GALLINARUM DNA-RELAXING ENZYME TO INVESTIGATE THE RELATIONSHIP BETWEEN THE NUMBER OF SUPERHELICAL TURNS AND THE MOLECULAR-WEIGHT IN A NEGATIVELY TWISTED DNA

Covalently closed-circular, superhelical DNAs, including viral DNAs, bacterial plasmid DNAs, and bacteriophage replicative-form DNA, were treated with a small amount of Haemo-philus gallinarum DNA-relaxing enzyme to generate incompletely relaxed DNA molecules. Each sample consisted of a set of closed-circular DNA molecules differing by one turn in their number of superhelical turns. The DNA samples were analyzed by agarose gel electro-phoresis under conditions such that the electrophoretic mobility was a function of the number of turns. The numbers of superhelical turns (at 37°C in 20 mM Tris-HCl (pH 7.5)-5 mM [MgCl2) in the DNAs of pSC101 (5.8 megadaltons), Colicin E1 (4.2 megadaltons), pMR4 (4.0 megadaltons; recombinant between pBR322 and λDNA fragment), φX174 replicative-form (RF) I, Simian virus 40 (SV40), and polyoma virus (3.4-3.6 megadaltons each), and λdv021 (2.05 megadaltons) were estimated to be 36, 27, 23-24, 20-21, 20-21, 20-21, and 11-13, respectively. It appears that the number of superhelical turns is mainly a function of the molecular weight of the DNA, at least in the substrates tested here. © 1979 by The Journal of Biochemistry.