Harpst J A, Dawson J R
Department of Biochemistry, Case Western Reserve University, Cleveland, Ohio 44106.
Biophys J. 1989 Jun;55(6):1237-49. doi: 10.1016/S0006-3495(89)82919-4.
Static light scattering measurements have been made at angles as low as 8 degrees on whole, half, and quarter molecules of native, T2 bacteriophage DNA in 0.195 M Na+. The fragments were obtained by high-speed stirring of the native DNA, and fractionated on methylated-albumin-kieselguhr columns. Accompanying measurements of sedimentation coefficients and intrinsic viscosities were made. Because linear extrapolations of light scattering data above 8 degrees for these samples were suspect, the measurements were analyzed by fitting curves calculated from the theory of wormlike coils to experimental curves at c = 0. Results showed that the excluded volume parameter, epsilon, must be used in analyzing the scattering curves; a reasonable value of epsilon was 0.08, in agreement with that found for T7 DNA (Harpst, J. A. 1980. Biophys. Chem. 11:295-302). The persistence length of all three DNAs in this paper was 50 +/- 5 nm, showed no dependence on molecular weight, but was somewhat below that reported previously for T7 DNA (60 nm). Theoretical curves calculated with the preceding parameters had a clear upward curvature in scattering envelopes below 8 degrees for quarter and half molecules, but such curvature was minimal for whole T2 DNA, so that linear extrapolations of experimental data above 8 degrees gave a molecular weight and root-mean-square radius which were nearly the same as those from theory. The molecular weight and radius for whole T2, derived from the comparison of theory and experiment, were 115 X 10(6) and 1,224 nm, respectively. The measurements on T2 DNA were clearly at the upper limit of current techniques.
在0.195 M Na⁺中,对天然T2噬菌体DNA的完整、半分子和四分之一分子进行了低至8度角的静态光散射测量。这些片段是通过对天然DNA进行高速搅拌获得的,并在甲基化白蛋白 - 硅藻土柱上进行分级分离。同时进行了沉降系数和特性粘度的测量。由于这些样品在8度以上的光散射数据的线性外推值得怀疑,因此通过将由蠕虫状线圈理论计算的曲线与c = 0时的实验曲线拟合来分析测量数据。结果表明,在分析散射曲线时必须使用排除体积参数ε;ε的合理值为0.08,与T7 DNA的结果一致(哈普斯特,J. A. 1980.生物物理化学. 11:295 - 302)。本文中所有三种DNA的持久长度为50±5 nm,与分子量无关,但略低于先前报道的T7 DNA的持久长度(60 nm)。用上述参数计算的理论曲线在低于8度时,对于四分之一和半分子的散射包络有明显的向上弯曲,但对于完整的T2 DNA,这种弯曲最小,因此8度以上实验数据的线性外推得到的分子量和均方根半径与理论值几乎相同。通过理论与实验比较得出的完整T2的分子量和半径分别为115×10⁶和1224 nm。对T2 DNA的测量显然处于当前技术的上限。