Mendz G L, Miller D J, Ralston G B
School of Biochemistry and Molecular Genetics, University of New South Wales, Sydney, Australia.
Eur Biophys J. 1995;24(1):39-53. doi: 10.1007/BF00216829.
The stoichiometry of palmitoyllysophosphatidylcholine/myelin basic protein (PLPC/MBP) complexes, the location of the protein in the lysolipid micelles, and the conformational changes occurring in the basic protein and peptides derived from it upon interaction with lysolecithin micelles were investigated by circular dichroic spectropolarimetry, ultracentrifugation, electron paramagnetic resonance (EPR) and 31P, 13C, and 1H nuclear magnetic resonance spectroscopy (NMR), and electron magnetic resonance spectroscopy (NMR), and electron microscopy. Ultracentrifugation measurements indicated that well-defined complexes were formed by the association of one protein molecule with approximately 141 lysolipid molecules. Small-angle X-ray scattering data indicated that the PLPC/MBP complexes form particles with a radius of gyration of 3.8 nm. EPR spectral parameters of the spin labels 5-, and 16-doxylstearate incorporated into lysolecithin/basic protein aggregates, and 13C- and 1H-NMR relaxation times of PLPC indicated that the addition of the protein did not affect the environment and location of the labels and the organization of the lysolipid micelles. The data suggested that MBP lies primarily near the surface of the micelles, with segments penetrating beyond the interfacial region into the hydrophobic interior, but without any part of the protein being protected against rapid exchange of its amide groups with the aqueous environment. The basic protein acquired about 20% alpha-helix when bound to lysolipid micelles. Circular dichroic spectra of sequential peptides derived by cleavage of the protein revealed the formation of alpha-helical regions in the association with lysolecithin. Specific residues in myelin basic protein that participated in binding to the micelles were identified from magnetic resonance data on changes in the chemical shifts and intensities of assigned resonances, and line broadening of peaks by fatty acid spin-labels incorporated into the micelles.
通过圆二色光谱极谱法、超速离心、电子顺磁共振（EPR）以及31P、13C和1H核磁共振光谱（NMR）和电子显微镜，研究了棕榈酰溶血磷脂酰胆碱/髓鞘碱性蛋白（PLPC/MBP）复合物的化学计量、蛋白质在溶血脂质微团中的位置，以及碱性蛋白及其衍生肽与溶血卵磷脂微团相互作用时发生的构象变化。超速离心测量表明，一个蛋白质分子与大约141个溶血脂质分子缔合形成了明确的复合物。小角X射线散射数据表明，PLPC/MBP复合物形成了回转半径为3.8nm的颗粒。掺入溶血卵磷脂/碱性蛋白聚集体中的自旋标记物5-和16-硬脂酰氧基硬脂酸酯的EPR光谱参数，以及PLPC的13C和1H-NMR弛豫时间表明，蛋白质的加入不影响标记物的环境和位置以及溶血脂质微团的组织。数据表明，MBP主要位于微团表面附近，部分片段穿透界面区域进入疏水内部，但蛋白质没有任何部分受到保护，其酰胺基团不会与水环境快速交换。碱性蛋白与溶血脂质微团结合时获得了约20%的α-螺旋。通过蛋白质裂解得到的连续肽段的圆二色光谱揭示了与溶血卵磷脂缔合时α-螺旋区域的形成。从关于指定共振的化学位移和强度变化以及掺入微团中的脂肪酸自旋标记物导致的峰线展宽的磁共振数据中，鉴定出了髓鞘碱性蛋白中参与与微团结合的特定残基。
