Kageyama-Yahara Natsuko, Riezman Howard
Department of Biochemistry, University of Geneva, Sciences II, 30, quai E. Ansermet, CH-1211 Geneva 4, Switzerland.
Biochem J. 2006 Sep 15;398(3):585-93. doi: 10.1042/BJ20060697.
Ceramide plays a crucial role as a basic building block of sphingolipids, but also as a signalling molecule mediating cell-fate decisions. Three genes, LAG1, LAC1 and LIP1, have been shown to be required for ceramide synthase activity in Saccharomyces cerevisiae [Guillas, Kirchman, Chuard, Pfefferli, Jiang, Jazwinski and Conzelman (2001) EMBO J. 20, 2655-2665; Schorling, Vallee, Barz, Reizman and Oesterhelt (2001) Mol. Biol. Cell 12, 3417-3427; Vallee and Riezman (2005) EMBO J. 24, 730-741]. In the present study, the topology of the Lag1p and Lac1p subunits was investigated. The N- and C-termini of the proteins are in the cytoplasm and eight putative membrane-spanning domains were identified in Lag1p and Lac1p by insertion of glycosylation and factor Xa cleavage sites at various positions. The conserved Lag motif, potentially containing the active site, is most likely embedded in the membrane. We also present evidence that histidine and aspartic acid residues in the Lag motif are essential for the function of Lag1p in vivo.
神经酰胺作为鞘脂的基本组成部分发挥着关键作用，同时也是介导细胞命运决定的信号分子。三个基因，LAG1、LAC1和LIP1，已被证明是酿酒酵母中神经酰胺合酶活性所必需的[Guillas、Kirchman、Chuard、Pfefferli、Jiang、Jazwinski和Conzelman（2001年）《欧洲分子生物学组织杂志》20，2655 - 2665；Schorling、Vallee、Barz、Reizman和Oesterhelt（2001年）《分子生物学细胞》12，3417 - 3427；Vallee和Riezman（2005年）《欧洲分子生物学组织杂志》24，730 - 741]。在本研究中，对Lag1p和Lac1p亚基的拓扑结构进行了研究。蛋白质的N端和C端位于细胞质中，通过在不同位置插入糖基化和因子Xa切割位点，在Lag1p和Lac1p中鉴定出八个假定的跨膜结构域。保守的Lag基序可能包含活性位点，很可能嵌入膜中。我们还提供证据表明，Lag基序中的组氨酸和天冬氨酸残基对于Lag1p在体内的功能至关重要。
