Stewart R J, Boggs J M
Department of Clinical Biochemistry, University of Toronto, Ontario, Canada.
Biochemistry. 1993 Jun 1;32(21):5605-14. doi: 10.1021/bi00072a016.
Factors which influence the accessibility, or exposure, of the carbohydrate head group of the glycolipid galactosylceramide (GalCer) at the membrane surface have been examined in lipid model membranes using the technique of galactose oxidase-tritiated sodium borohydride labeling. Both the ceramide composition of GalCer and the lipid composition of its membrane environment were varied. We have shown that GalCer is oxidized in a membrane environment, by purification of the labeled galactosyl moiety of the glycolipid by high-performance anion exchange chromatography. Using semisynthetic molecular species of GalCer with acyl chain lengths ranging from 16 to 26 carbons, incorporated into liposome membranes of egg phosphatidylcholine (PC), and reverse-phase HPLC separation of mixtures of the molecular species, we have shown that increasing the fatty acid chain length of GalCer increases its oxidation by galactose oxidase. In addition, the degree of oxidation is reduced when the fatty acid chain of GalCer is hydroxylated. GalCer incorporated into liposomes containing synthetic species of PC with different fatty acid chain lengths (together with cholesterol) was oxidized less as the PC acyl chain length, and hence the bilayer thickness, was increased. The oxidation of GalCer in liposomes composed of sphingomyelin/cholesterol was reduced compared to its oxidation in PC liposomes. Furthermore, changes in the fatty acid chain length of GalCer had no effect on its oxidation in sphingomyelin liposomes. These findings indicate that the ceramide composition and lipid membrane environment can influence the exposure of the lipid carbohydrate, and hence, they could modulate the receptor activity of glycolipids at the membrane surface.
利用半乳糖氧化酶-氚化硼氢化钠标记技术，在脂质模型膜中研究了影响糖脂半乳糖神经酰胺（GalCer）碳水化合物头部基团在膜表面可及性或暴露程度的因素。GalCer的神经酰胺组成及其膜环境的脂质组成均有所变化。通过高效阴离子交换色谱法纯化糖脂的标记半乳糖部分，我们证明了GalCer在膜环境中会被氧化。使用酰基链长度在16至26个碳之间的GalCer半合成分子种类，将其掺入鸡蛋磷脂酰胆碱（PC）的脂质体膜中，并通过反相高效液相色谱法分离分子种类的混合物，我们发现增加GalCer的脂肪酸链长度会增加其被半乳糖氧化酶氧化的程度。此外，当GalCer的脂肪酸链被羟基化时，氧化程度会降低。掺入含有不同脂肪酸链长度（以及胆固醇）的PC合成种类的脂质体中的GalCer，随着PC酰基链长度增加，进而双层厚度增加，其氧化程度降低。与在PC脂质体中的氧化相比，GalCer在由鞘磷脂/胆固醇组成的脂质体中的氧化程度降低。此外，GalCer脂肪酸链长度的变化对其在鞘磷脂脂质体中的氧化没有影响。这些发现表明，神经酰胺组成和脂质膜环境可以影响脂质碳水化合物的暴露，因此，它们可以调节糖脂在膜表面的受体活性。
