Demeler Borries, van Holde Kensal E
Department of Biochemistry, University of Texas Health Science Center at San Antonio, San Antonio, TX 78229, USA.
Anal Biochem. 2004 Dec 15;335(2):279-88. doi: 10.1016/j.ab.2004.08.039.
This article discusses several improvements to the van Holde-Weischet (vHW) method [Biopolymers 17 (1978) 1387] that address its capability to deal with sedimentation coefficient distributions spanning a large range of s values. The method presented here allows the inclusion of scans early and late in the experiment that ordinarily would need to be excluded from the analysis due to ultracentrifuge cell end effects. Scans late in the experiment are compromised by the loss of a defined plateau region and by back-diffusion from the bottom of the cell. Early scans involve partial boundaries that have not fully cleared the meniscus. In addition, a major refinement of the algorithm for determining the boundary fractions is introduced, taking into account different degrees of radial dilution for different species in the system. The method retains its desirable model-independent properties (the analysis of sedimentation data does not require prior knowledge of a user-imposed model or range of sedimentation coefficients) and reports diffusion-corrected s value distributions, which can be presented either in a histogram format or the traditional integral distribution format. Data analyzed with the traditional vHW method are compared with those of the improved method to demonstrate the benefit from the added information in the analysis.
本文讨论了对范霍尔德 - 魏谢特(vHW)方法[《生物聚合物》17(1978)1387]的若干改进,这些改进解决了该方法处理跨越较大s值范围的沉降系数分布的能力问题。本文提出的方法允许纳入实验早期和晚期的扫描数据,这些扫描数据通常因超速离心池的端部效应而需要从分析中排除。实验晚期的扫描数据受到明确平台区域的损失以及从池底部的反向扩散的影响。早期扫描涉及尚未完全清除弯月面的部分边界。此外,引入了用于确定边界分数的算法的一项重大改进,考虑了系统中不同物种的不同程度的径向稀释。该方法保留了其理想的与模型无关的特性(沉降数据分析不需要用户强加的模型或沉降系数范围的先验知识),并报告经扩散校正的s值分布,该分布可以以直方图格式或传统的积分分布格式呈现。将用传统vHW方法分析的数据与改进方法的数据进行比较,以证明分析中添加信息的益处。