McLaurin J, Ackerley C A, Moscarello M A
Department of Biochemistry, Hospital for Sick Children, Toronto, Canada.
J Neurosci Res. 1993 Aug 15;35(6):618-28. doi: 10.1002/jnr.490350605.
The myelin basic protein (MBPs) represent a family of proteins (charge isomers) which account for 35% of the total myelin protein. Localization studies have been inconclusive because MBP is not a single protein. Antibodies obtained by injection of MBP into animals recognized all members of the MBP family. In the studies reported here, we have fractionated the MBPs into specific components or charge isomers. One of these which contains citrulline accounts for about 20% of the total MBP. We report the localization of this single MBP to the intraperiod line of myelin by immunoelectron microscopy. For these studies several specific antibodies were used including antibodies raised against total MBP, specific MBP peptides, and against a tetracitrulline peptide. This latter antibody was specific for component 8 (C-8) of MBP. Since C-8 is the only MBP which contains citrulline it was used to localize this particular form of MBP principally to the intraperiod line by immunogold electron microscopy, while antibody against total MBP (consisting of all charge isomers C-1-->C-8) labelled both the major dense line and the intraperiod line. When the anti-citrulline antibody was used with a 3 nm gold conjugated Fab fragments prepared from the secondary antibody, 66.5% of the gold particles were localized to the intraperiod line, while 11.2% of gold particles were localized to the major dense line. On the other hand, with the monoclonal anti-MBP antibodies reactive with residues 69-74, 59.4% of the gold particles were localized to the major dense line and 23.6% of gold particles at the intraperiod line. Other supporting evidence includes increased labelling of myelin by 125I labelled anti-citrulline IgG when isolated myelin was swollen, a process known to take place at the intraperiod line. Gold particles were demonstrated at the intraperiod line in swollen and recompacted myelin. C-8 was shown to associate preferentially with lipids asymmetrically localized to the intraperiod line.
髓鞘碱性蛋白（MBP）是一类蛋白质（电荷异构体）家族，占髓鞘总蛋白的35%。由于MBP不是单一蛋白质，定位研究尚无定论。通过将MBP注射到动物体内获得的抗体可识别MBP家族的所有成员。在本文报道的研究中，我们已将MBP分离为特定成分或电荷异构体。其中一种含瓜氨酸的异构体约占总MBP的20%。我们通过免疫电子显微镜报告了这种单一MBP在髓鞘内节线的定位。对于这些研究，使用了几种特异性抗体，包括针对总MBP、特定MBP肽段以及针对四瓜氨酸肽段产生的抗体。后一种抗体对MBP的成分8（C-8）具有特异性。由于C-8是唯一含瓜氨酸的MBP，因此通过免疫金电子显微镜主要将这种特定形式的MBP定位到内节线，而针对总MBP（由所有电荷异构体C-1至C-8组成）的抗体则标记了主致密线和内节线。当抗瓜氨酸抗体与由二抗制备的3nm金偶联Fab片段一起使用时，66.5%的金颗粒定位于内节线，而11.2%的金颗粒定位于主致密线。另一方面，与69 - 74位残基反应的单克隆抗MBP抗体，59.4%的金颗粒定位于主致密线，23.6%的金颗粒定位于内节线。其他支持证据包括当分离的髓鞘肿胀时，125I标记的抗瓜氨酸IgG对髓鞘的标记增加，这一过程已知发生在内节线。在肿胀和重新压实的髓鞘中，在内节线处发现了金颗粒。已表明C-8优先与不对称定位于内节线的脂质结合。
