Libich David S, Harauz George
Department of Molecular and Cellular Biology, and Biophysics Interdepartmental Group, University of Guelph, 50 Stone Road East, N1G 2W1, Guelph, ON, Canada.
Eur Biophys J. 2008 Jul;37(6):1015-29. doi: 10.1007/s00249-008-0334-8. Epub 2008 May 1.
The stability and secondary structure propensity of recombinant murine 18.5 kDa myelin basic protein (rmMBP, 176 residues) was assessed using circular dichroic and nuclear magnetic resonance spectroscopy (1H-15N HSQC experiments) to determine the optimal sample conditions for further NMR studies (i.e., resonance assignments and protein-protein interactions). Six solvent conditions were selected based on their ability to stabilise the protein, and their tractability to currently standard solution NMR methodology. Selected solvent conditions were further characterised as functions of concentration, temperature, and pH. The results of these trials indicated that 30% TFE-d2 in H2O (v/v), pH 6.5 at 300 K, and 100 mM KCl, pH 6.5 at 277 K were the best conditions to use for future solution NMR studies of MBP. Micelles of DPC were found to be inappropriate for backbone resonance assignments of rmMBP in this instance.
利用圆二色光谱和核磁共振波谱（1H-15N HSQC实验）评估重组小鼠18.5 kDa髓鞘碱性蛋白（rmMBP，176个残基）的稳定性和二级结构倾向，以确定用于进一步核磁共振研究（即共振归属和蛋白质-蛋白质相互作用）的最佳样品条件。基于其稳定蛋白质的能力及其对当前标准溶液核磁共振方法的易处理性，选择了六种溶剂条件。所选溶剂条件进一步表征为浓度、温度和pH的函数。这些试验结果表明，30%（体积/体积）TFE-d2溶于H2O、300 K时pH 6.5以及100 mM KCl、277 K时pH 6.5是未来用于MBP溶液核磁共振研究的最佳条件。在这种情况下，发现DPC胶束不适用于rmMBP的主链共振归属。
