High-Yield Preparation of Oligomeric C-Type DNA Toroids and Their Characterization by Cryoelectron Microscopy

Abstract

The quantitative and reversible compaction of open circular plasmid DNA (7676 bp) into toroids containing one to 19 molecules by sequential treatment with spermine and an excess of uranyl acetate is reported. The toroidal DNA structure was proven by cryoelectron microscopy. Linearized and supercoiled variants of the DNA also gave toroids under these conditions, but yields were significantly lower. In the presence of spermine alone no toroids were found. Open circular plasmid B-DNA helix was converted into the C-type helical form upon compaction as was shown by CD spectroscopy (negative peak at 255 nm) and electron microscopy (1.8-nm interduplex distance instead of 2.9 nm). Addition of uranyl salt to the DNA−spermine complexes resulted in the formation of netlike assemblies which further compacted to give toroids. A model containing a hexagonal arrangement of DNA strands with extensive strand crossings is proposed. Curvature and thus toroid formation is thought to be induced by the hydrophobic DNA coating ...