LIGHT-SCATTERING-STUDIES OF SUPERCOILED AND NICKED DNA

Static and dynamic light scattering measurements are made of solutions of pGem1a plasmids (3730 base pairs) in the relaxed circular (nicked) and supercoiled forms. The static structure factor and the spectrum o f decay modes in the autocorrelation function were examined in order t o determine the salient differences between the behaviors of nicked DN A and supercoiled DNA. The concentrations studied are within the dilut e regime, which is to say that the structure and dynamics of an isolat ed DNA molecule were probed. Static light scattering measurements yiel ded estimates for the molecular weight M, second virial coefficient A( 2), and radius of gyration R(G). For the nicked DNA, M = (2.8 +/- 0.4) x 10(6) g/mol, A(2) = (0.9 +/- 0.2) x 10(-3) mol cm(3)/g(2), and R(G) = 90 +/- 3 nm were obtained. For the supercoiled DNA, M = (2.5 +/- 0. 4) x 10(6) g/mol, A(2) = (1.2 +/- 0.2) x 10(-3) mol cm(3)/g(2), and R( G) = 82 +/- 2.5 nm were obtained. The static structure factors for the nicked and supercoiled DNA were found to superpose when they were sca led by the radius of gyration. The intrinsic stiffness of DNA was evid ent in the static light scattering data. Homodyne intensity autocorrel ation functions were collected by both DNAs at several angles, or scat tering vectors. At the smallest scattering vectors the probe size was comparable to the longest intramolecular distance, while at the larges t scattering vectors the probe size was smaller than the persistence l ength of the DNA. Values of the self-diffusion coefficients D were obt ained from the low-angle data. For the nicked DNA, D = (2.9 +/- 0.3) x 10(-8) cm(2)/s. The contribution to the correlation function from the internal dynamics of the DNA was seen to result in a strictly bimodal decay function. The rates of the faster model Gamma(int), reached pla teau values at low angles. For the nicked DNA, Gamma(int) = 2500 +/- 5 00 s(-1), and for the supercoiled DNA, Gamma(int) = 5000 +/- 500 s(-1) . These rates correspond to the slowest internal relaxation modes of t he DNAs. The dependence of the relaxation rates on scattering vector w as monitored with the aid of cumulants analysis and compared with theo retical predictions for the semiflexible ring molecule. The internal m ode rates and the dependence of the cumulants moments reflected the di fference between the nicked DNA and the supercoiled DNA dynamical beha vior. The supercoiled DNA behavior seen here indicates that conformati onal dynamics might play a larger role in DNA behavior than is suggest ed by the notion of a branched interwound structure. (C) 1996 John Wil ey & Sons, Inc.