Wang T Y, Silvius J R
Department of Biochemistry, McGill University, Montréal, Québec H3G 1Y6, Canada.
Biophys J. 2000 Sep;79(3):1478-89. doi: 10.1016/S0006-3495(00)76399-5.
Two fluorescence-based approaches have been applied to examine the differential partitioning of fluorescent phospho- and sphingolipid molecules into sphingolipid-enriched domains modeling membrane "lipid rafts." Fluorescence-quenching measurements reveal that N-(diphenylhexatrienyl)propionyl- (DPH3:0-)-labeled gluco- and galactocerebroside partition into sphingolipid-enriched domains in sphingolipid/phosphatidylcholine/cholesterol bilayers with substantially higher affinity than do analogous sphingomyelin, ceramide, or phosphatidylcholine molecules. By contrast, the affinity of sphingomyelin and ceramide for such domains is only marginally greater than that of a phosphatidylcholine with similar hydrocarbon chains. By using direct measurements of molecular partitioning between vesicles of different compositions, we show that the relative affinities of different C(6)-NBD- and C(5)-Bodipy-labeled sphingolipids for sphingolipid-enriched domains are quantitatively, and in most circumstances even qualitatively, quite different from those found for species whose N-acyl chains more closely resemble the long saturated chains of cellular sphingolipids. These findings lend support in principle to previous suggestions that differential partitioning of different sphingolipids into "raft" domains could contribute to the differential trafficking of these species in eukaryotic cells. However, our findings also indicate that short-chain sphingolipid probes previously used to examine this phenomenon are in general ill-suited for such applications.
两种基于荧光的方法已被用于研究荧光磷脂和鞘脂分子在模拟膜“脂筏”的富含鞘脂结构域中的差异分配。荧光猝灭测量表明，N-（二苯基己三烯基）丙酰基-（DPH3:0-）标记的葡萄糖脑苷脂和半乳糖脑苷脂在鞘脂/磷脂酰胆碱/胆固醇双层膜中分配到富含鞘脂的结构域时，其亲和力远高于类似的鞘磷脂、神经酰胺或磷脂酰胆碱分子。相比之下，鞘磷脂和神经酰胺对这些结构域的亲和力仅略高于具有相似烃链的磷脂酰胆碱。通过直接测量不同组成囊泡之间的分子分配，我们发现不同C（6）-NBD-和C（5）-Bodipy标记的鞘脂对富含鞘脂结构域的相对亲和力在数量上，而且在大多数情况下甚至在质量上，与那些N-酰基链更类似于细胞鞘脂长饱和链的物种的情况有很大不同。这些发现原则上支持了先前的观点，即不同鞘脂在“筏”结构域中的差异分配可能有助于这些物种在真核细胞中的差异运输。然而，我们的发现也表明，先前用于研究这一现象的短链鞘脂探针通常不适用于此类应用。
