The GPI transamidase complex of Saccharomyces cerevisiae contains Gaa1p, Gpi8p, and Gpi16p
The GPI transamidase complex of Saccharomyces cerevisiae contains Gaa1p, Gpi8p, and Gpi16p
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DOI：
10.1091/mbc.12.10.3295
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发表时间：
2001-10-01
影响因子：
3.3
通讯作者：
Conzelmann, A
中科院分区：
文献类型：
--
作者：
Fraering, P;Imhof, I;Conzelmann, A
Gpi8p和Gaa1p是GPI转氨酶的重要组分，GPI转氨酶将糖基磷脂酰肌醇（GPI）添加到新合成的蛋白质中。在1.5%洋地黄苷溶液中溶解后，用蓝色原生PAGE分离，Gpi8p存在于430-650-kDa蛋白复合物中。这些复合物可以亲和纯化，由Gaa1p、Gpi8p和Gpi16p （YHR188c）组成。Gpi16p是必需的n -糖基化跨膜糖蛋白。它的主体位于内质网的管腔侧，具有单个c端跨膜结构域和一个小的c端细胞质延伸，具有内质网检索基序。Gpi16p的缺失导致完整的GPI脂质CP2和未加工的GPI前体蛋白的积累。Gpi8p和Gpi16p是不稳定的，如果它们中的任何一个被耗尽。同样，当Gpi8p过表达时，它大部分停留在430-650-kDa转氨酶复合物之外，并且不稳定。Gpi8p的过表达不能弥补Gpi16p的缺失。Gpi16p的同源物存在于所有真核生物中。转氨酶复合物分别与内质网插入所需的Sec61p复合物和GPI蛋白的n -糖基化所需的低聚糖转移酶复合物无关。当GPI前体蛋白或GPI脂质被耗尽时，转氨酶复合物保持完整。
Gpi8p and Gaa1p are essential components of the GPI transamidase that adds glycosylphosphatidylinositols (GPIs) to newly synthesized proteins. After solubilization in 1.5% digitonin and separation by blue native PAGE, Gpi8p is found in 430-650-kDa protein complexes. These complexes can be affinity purified and are shown to consist of Gaa1p, Gpi8p, and Gpi16p (YHR188c). Gpi16p is an essential N-glycosylated transmembrane glycoprotein. Its bulk resides on the lumenal side of the ER, and it has a single C-terminal transmembrane domain and a small C-terminal, cytosolic extension with an ER retrieval motif. Depletion of Gpi16p results in the accumulation of the complete GPI lipid CP2 and of unprocessed GPI precursor proteins. Gpi8p and Gpi16p are unstable if either of them is removed by depletion. Similarly, when Gpi8p is overexpressed, it largely remains outside the 430-650-kDa transamidase complex and is unstable. Overexpression of Gpi8p cannot compensate for the lack of Gpi16p. Homologues of Gpi16p are found in all eucaryotes. The transamidase complex is not associated with the Sec61p complex and oligosaccharyltransferase complex required for ER insertion and N-glycosylation of GPI proteins, respectively. When GPI precursor proteins or GPI lipids are depleted, the transamidase complex remains intact.