Mamelak D, Mylvaganam M, Tanahashi E, Ito H, Ishida H, Kiso M, Lingwood C
Research Institute, Hospital for Sick Children, 555 University Avenue, Toronto, Ont., Canada M5G 1X8.
Carbohydr Res. 2001 Sep 28;335(2):91-100. doi: 10.1016/s0008-6215(01)00209-9.
3'-Sulfogalactolipids(SGLs), sulfogalactosyl ceramide (SGC), and sulfogalactoglycerolipid (SGG) bind to the N-terminal ATPase-containing domain of members of the heat shock protein 70 family. We have probed this binding specificity using a series of synthetic positional sulfated or phosphorylated glycolipid analogues, containing either a long-chain bisalkyl hydrocarbon-2-(tetradecyl)hexadecane (B30) or C(18) ceramide (SGC(18)) backbone. By TLC overlay and receptor ELISA, recombinant hsc70 bound ceramide-based glycoconjugates having 3'- or 4'-sulfogalactose glycone moieties and the 4'-sulfogalactose positional isomer conjugated to B30. Hsc70 binding was significantly decreased to the 3'-sulfogalactose conjugated to the long-chain branched alkane. 3'-Sulfoglucose conjugated to B30 was not bound, nor were similarly conjugated di-, tri-, and tetra-sulfated or phosphorylated galactolipids. These results highlight the importance of the position, rather than the number of sulfate esters within the galactose ring. This binding selectivity was shared by the sea urchin hsp70-related sperm receptor. A 3'-SGC-based soluble inhibitor, in which the acyl chain was replaced with an adamantyl group, inhibited binding of hsc70 to both 3'- and 4'-SGC species with an IC(50) of 50 and 75 microM, respectively, indicating a shared sulfogalactose binding site. These studies demonstrate the highly specific nature of hsc70/SGL binding and show, for the first time, that the lipid aglycone can alter the substitution position requirement for glycolipid recognition.
3'-硫酸半乳糖脂（SGLs）、硫酸半乳糖神经酰胺（SGC）和硫酸半乳糖甘油脂（SGG）与热休克蛋白70家族成员含N端ATP酶的结构域结合。我们使用了一系列合成的位置硫酸化或磷酸化糖脂类似物来探究这种结合特异性，这些类似物含有长链双烷基烃-2-（十四烷基）十六烷（B30）或C（18）神经酰胺（SGC（18））骨架。通过薄层层析覆盖法和受体酶联免疫吸附测定，重组hsc70与具有3'-或4'-硫酸半乳糖糖基部分以及与B30共轭的4'-硫酸半乳糖位置异构体的基于神经酰胺的糖缀合物结合。hsc70与与长链支链烷烃共轭的3'-硫酸半乳糖的结合显著减少。与B30共轭的3'-葡萄糖未被结合，类似共轭的二、三、四硫酸化或磷酸化半乳糖脂也未被结合。这些结果突出了半乳糖环内硫酸酯位置而非数量的重要性。这种结合选择性也存在于海胆hsp70相关的精子受体中。一种基于3'-SGC的可溶性抑制剂，其中酰基链被金刚烷基取代，分别以50和75 microM的IC（50）抑制hsc70与3'-和4'-SGC种类的结合，表明存在共同的硫酸半乳糖结合位点。这些研究证明了hsc70/SGL结合的高度特异性，并首次表明脂质苷元可改变糖脂识别的取代位置要求。
