Polylysine-coated mica can be used to observe systematic changes in the supercoiled DNA conformation by scanning force microscopy in solution.
The conformations of supercoiled (sc) DNA and linear DNA bound to polylysine (PL)-coated mica were investigated by scanning force microscopy (SFM) in solution. From the polymer statistical analysis of linear DNA, we could distinguish between re-arrangements or trapping of the DNA on the surface. Conditions of re-arrangements to an almost equilibrated state can be achieved at appropriate PL surface concentrations. We could show that the ability of re-arrangements depends on the salt concentration of the adsorption/imaging buffer. Comparing the statistical analysis of the linear DNA with SFM images of scDNA suggested that irregular scDNA conformations are formed under conditions of trapping, whereas plectonemic structures are favoured under conditions of surface re-arrangements. Salt-dependent changes in the scDNA conformation over the range of 10-100 mM NaCl, as characterised by the parameters writhe and the superhelix radius r, are observable only under conditions that enable surface re-arrangements. The measured values of writhe suggest that the scDNA loses approximately one-half of the supercoils during the binding to the surface. At the same time r increases systematically with decreasing writhe, thus the scDNA topology remains determined by the constraints on supercoiling during the binding to PL-coated mica.
Bussiek
Malte
M
German Cancer Research Center, Department of Biophysics of Macromolecules, Im Neuenheimer Feld 580, D-69120 Heidelberg, Germany.
Mücke
Norbert
N
Langowski
Jörg
J
D000538:Aluminum Silicates / Q000737:chemistry*; D004278:DNA, Superhelical / Q000737:chemistry* / Q000648:ultrastructure; D018625:Microscopy, Atomic Force; D009690:Nucleic Acid Conformation / Q000187:drug effects; D011107:Polylysine / Q000737:chemistry*; D012965:Sodium Chloride / Q000494:pharmacology; D013499:Surface Properties
D000538:Aluminum Silicates; D004278:DNA, Superhelical; D011107:Polylysine; D012965:Sodium Chloride; C011934:mica