Acquisition of an Optima XL-I Analytical Ultracentrifuge

9977029摘要本项目涉及佛蒙特大学分子生理与生物物理系购买和安装一台最先进的分析式超离心机。该仪器代表了最新发展的技术，完全计算机化，提供方便和快速的数据采集和操作。计算机程序可用于执行许多计算，以确定分子量，沉降和扩散系数以及大分子的均匀性或多分散性程度。这项新技术最重要的应用之一将是能够定量分析执行大多数细胞功能的复杂大分子组合的缔合-解离平衡。随着分子生物学的快速发展以及在细菌和杆状病毒表达系统中创造无限新蛋白质的前所未有的能力，迫切需要通过精确的物理化学技术来表征野生型和突变蛋白。分析式超离心机仍然是这类研究的首选工具。主要用户的研究致力于了解大分子及其组装的结构和功能，特别是那些涉及肌肉收缩和非肌肉运动的大分子。大多数项目将使用仪器来研究分子马达，肌球蛋白及其与肌动蛋白的相互作用的某些方面；轻链异构体在心肌肌球蛋白中的作用钙调蛋白对非肌凝蛋白的调控及平滑肌肌凝蛋白的组装特性单个肌球蛋白分子产生的力和运动；肌动球蛋白界面的分子相互作用；果蝇肌球蛋白调控轻链在振荡发电中的作用心力衰竭中的细丝调节。其中几个项目将使用仪器来表征表达蛋白的均匀性，这是光学陷阱中单分子力学必不可少的特性；还有x射线晶体学。此外，该仪器将成为研究生和博士后研究员以及本科生的优秀教学工具。很少有仪器能像这样生动地展示环境条件对生物大分子的大小、形状和相互作用特性的影响。

9977029Abstract This project involves the acquisition and installation of a state-of-the-art-analytical ultracentrifuge in the Department of Molecular Physiology and Biophysics at the University of Vermont. This instrument represents newly developed technology, fully computerized to provide convenient and rapid data acquisition and manipulation. Computer programs are available to perform the many of the computations needed to determine molecular weights, sedimentation and diffusion coefficients and the degree of homogeneity or polydispersity of macromolecules. One of the most important applications of the new technology will be the ability to quantitatively analyze the association-dissociation equilibria of complex macromolecular assemblies that carry out most cellular functions. With the rapid advances in molecular biology and the unprecedented ability to create unlimited new proteins in bacterial and baculovirus expression systems, there is a compelling need to characterize wild type and mutant proteins by precise physical chemical techniques. The analytical ultracentrifuge remains the tool of choice for such studies. The research of the major users is dedicated to understanding the structure and function of macromolecules and their assemblies, in particular, those involved in muscle contraction and non-muscle motility. Most of the projects will use the instrumentation to study some aspect of the molecular motor, myosin, and its interactions with actin; the role of light chain isoforms in cardiac myosins; the regulation of non-muscle myosins by calmodulin, and assembly properties of smooth muscle myosin; forces and motions generated by single myosin molecules; molecular interactions at the actomyosin interface; role of Drosophila myosin regulatory light chain in oscillatory power generation; thin filament regulation in heart failure. Several of the projects will use the instrumentation to characterize the expressed proteins for homogeneity, a property essential for single molecule mechanics in optical traps; and for x-ray crystallography. In addition, this instrument will serve as an excellent teaching tool for graduate students and post-doctoral fellows as well as for undergraduates. Few instruments demonstrate as vividly as this the effects of environmental conditions on the size, shape and interaction properties of biological macromolecules.