The 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 Haemophilus 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 electrophoresis under conditions such that the electrophoretic mobility was a function of the number of turns. The numbers of superhelical turns (at 37 degrees 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 lambda DNA fragment), phi X174 replicative-form (RF) I, Simian virus 40 (SV40), and polyoma virus (3.4--3.6 megadaltons each), and lambda dv021 (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.
D002847:Chromatography, Agarose; D004264:DNA Topoisomerases, Type I; D004269:DNA, Bacterial / Q000032:analysis; D004270:DNA, Circular / Q000032:analysis; D004278:DNA, Superhelical / Q000032:analysis*; D004279:DNA, Viral / Q000032:analysis; D006190:Haemophilus / Q000201:enzymology*; D008970:Molecular Weight; D009690:Nucleic Acid Conformation
D004269:DNA, Bacterial; D004270:DNA, Circular; D004278:DNA, Superhelical; D004279:DNA, Viral; D004264:DNA Topoisomerases, Type I