Lipid trafficking controls endotoxin acylation in outer membranes of Escherichia coli
Lipid trafficking controls endotoxin acylation in outer membranes of Escherichia coli
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DOI：
10.1074/jbc.m404963200
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发表时间：
2004-10-22
影响因子：
4.8
通讯作者：
Bishop, RE
中科院分区：
文献类型：
--
作者：
Jia, WY;El Zoeiby, A;Bishop, RE
生物膜的生物发生取决于不同细胞区室之间膜脂的协调运输。细菌外膜酶 PagP 通过将棕榈酸酯链从磷脂转移到脂多糖的脂质 A（内毒素）成分，赋予宿主免疫防御抵抗力。 PagP 是一个八链反平行 β 桶，前面有一个 N 端两亲性 α 螺旋。活性位点位于 β 桶内部，并与含有脂多糖的外叶对齐，但磷脂底物通常仅限于不对称外膜的内叶。我们研究了 PagP 体内活性取决于磷脂异常迁移到外叶的可能性。我们发现，向大肠杆菌培养物中短暂添加毫摩尔 EDTA（据报道可以用磷脂取代一部分脂多糖）会快速诱导脂质 A 的棕榈酰化。虽然大肠杆菌 pagP 基因的表达是在 Mg2+ 限制期间通过 phoPQ 双组分信号转导途径诱导的，但 EDTA 诱导的脂质 A 棕榈酰化发生得比 pagP 诱导更快，并且与从头蛋白质合成无关。 EDTA 诱导的脂质 A 棕榈酰化需要功能性 MsbA，这是脂质转运至外膜所需的必需 ATP 结合盒转运蛋白。通过显示 α 螺旋缺失在体内活跃，排除了 PagP α 螺旋在磷脂易位到外叶中的潜在作用。 EDTA 和 Mg2+-EDTA 均不会刺激体外 PagP 活性。这些发现表明 PagP 在外膜中保持休眠状态，直到 Mg2+ 限制促进磷脂迁移到外叶中。
The biogenesis of biological membranes hinges on the coordinated trafficking of membrane lipids between distinct cellular compartments. The bacterial outer membrane enzyme PagP confers resistance to host immune defenses by transferring a palmitate chain from a phospholipid to the lipid A ( endotoxin) component of lipopolysaccharide. PagP is an eight-stranded antiparallel beta-barrel, preceded by an N-terminal amphipathic alpha-helix. The active site is localized inside the beta-barrel and is aligned with the lipopolysaccharide-containing outer leaflet, but the phospholipid substrates are normally restricted to the inner leaflet of the asymmetric outer membrane. We examined the possibility that PagP activity in vivo depends on the aberrant migration of phospholipids into the outer leaflet. We find that brief addition to Escherichia coli cultures of millimolar EDTA, which is reported to replace a fraction of lipopolysaccharide with phospholipids, rapidly induces palmitoylation of lipid A. Although expression of the E. coli pagP gene is induced during Mg2+ limitation by the phoPQ two-component signal transduction pathway, EDTA-induced lipid A palmitoylation occurs more rapidly than pagP induction and is independent of de novo protein synthesis. EDTA-induced lipid A palmitoylation requires functional MsbA, an essential ATP-binding cassette transporter needed for lipid transport to the outer membrane. A potential role for the PagP alpha-helix in phospholipid translocation to the outer leaflet was excluded by showing that alpha-helix deletions are active in vivo. Neither EDTA nor Mg2+-EDTA stimulate PagP activity in vitro. These findings suggest that PagP remains dormant in outer membranes until Mg2+ limitation promotes the migration of phospholipids into the outer leaflet.