Chan K F, Robb N D, Chen W H
Laboratory of Experimental Neuropathology, National Institute of Neurological Disorders and Stroke, National Institutes of Health, Bethesda, MD 20892.
J Neurosci Res. 1990 Apr;25(4):535-44. doi: 10.1002/jnr.490250410.
The structural characteristics of myelin basic protein (MBP) involved in protein-protein and protein-lipid interactions were investigated. Rabbit MBP could bind calmodulin (CaM) in the presence of Ca2+ to form a complex that remained undissociated in 8 M urea. However, no tight complex formation was observed when the divalent cation was absent. These results suggest that MBP may contain a hydrophobic domain similar to those in the other well-characterized CaM-binding proteins. The stoichiometry of calmodulin binding to MBP was approximately 1:1. Prior limited proteolysis of MBP with trypsin abolished the formation of the MBP-CaM complex, indicating that the entire MBP polypeptide may be involved in the recognition of the hydrophobic clefts in CaM. MBP also formed tight complexes with gangliosides, but the presence of Ca2+ was not required. Binding of gangliosides to MBP-CaM complex released CaM from the complex. The ganglioside-binding sites in MBP were determined after trisecting the protein at two glutamic acid residues with Staphylococcus aureus V8 protease. Subsequent binding studies revealed that a 9.5-kDa polypeptide, which may correspond to the NH2-terminal domain (residues 1-83) of MBP, had higher affinity for the binding of lucifer yellow CH-labeled GM1 than did the other two polypeptides, of apparent molecular mass (Mr) 5,500 and 4,500, respectively. Among the various proteins in purified guinea pig brain myelin, synaptosomes, and synaptosomal membranes, MBP was found to have the highest affinity in binding lucifer yellow CH-GM1.
研究了髓鞘碱性蛋白（MBP）参与蛋白质-蛋白质和蛋白质-脂质相互作用的结构特征。兔MBP在Ca2+存在下可与钙调蛋白（CaM）结合形成一种在8 M尿素中仍不解离的复合物。然而，当不存在二价阳离子时，未观察到紧密复合物的形成。这些结果表明，MBP可能含有一个与其他特征明确的CaM结合蛋白中类似的疏水结构域。CaM与MBP结合的化学计量比约为1:1。用胰蛋白酶对MBP进行预先有限的蛋白水解消除了MBP-CaM复合物的形成，表明整个MBP多肽可能参与了对CaM中疏水裂缝的识别。MBP还与神经节苷脂形成紧密复合物，但不需要Ca2+的存在。神经节苷脂与MBP-CaM复合物的结合会使CaM从复合物中释放出来。在用金黄色葡萄球菌V8蛋白酶在两个谷氨酸残基处将该蛋白三等分后，确定了MBP中的神经节苷脂结合位点。随后的结合研究表明，一条9.5 kDa的多肽，可能对应于MBP的NH2末端结构域（第1-83位氨基酸残基），对荧光素黄CH标记的GM1的结合亲和力高于另外两条表观分子量（Mr）分别为5500和4500的多肽。在纯化的豚鼠脑髓鞘、突触体和突触体膜中的各种蛋白质中，发现MBP对荧光素黄CH-GM1的结合亲和力最高。
