End-products diacylglycerol and ceramide modulate membrane fusion induced by a phospholipase C/sphingomyelinase from Pseudomonas aeruginosa
End-products diacylglycerol and ceramide modulate membrane fusion induced by a phospholipase C/sphingomyelinase from Pseudomonas aeruginosa
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DOI：
10.1016/j.bbamem.2009.10.017
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发表时间：
2010-01-01
影响因子：
3.4
通讯作者：
Goni, Felix M.
中科院分区：
文献类型：
--
作者：
Ibarguren, Maitane;Bomans, Paul H. H.;Goni, Felix M.
已在含有等摩尔比例的磷脂酰胆碱、鞘磷脂、磷脂酰乙醇胺和胆固醇的囊泡上测定了来自铜绿假单胞菌的磷脂酶C/鞘磷脂酶。酶活性改变双层化学成分，产生二酰基甘油 (DAG) 和神经酰胺 (Cer)。已经进行了酶活性、酶诱导的聚集和融合的测定。基于冷冻电镜观察，提出了该过程各个阶段的囊泡融合的超微结构证据。两种酶脂质终产物 DAG 和 Cer 对双层物理性质具有相反的影响；前者消除了横向相分离，而后者产生了新的凝胶相[Sot et al., FEBS Lett. 582, 3230-3236 (2008)]。将 DAG 或 Cer 或两者添加到脂质体混合物中会导致酶与双层的结合增加并减少水解的滞后时间。这两种脂质对酶活性也有不同的影响，DAG增强酶诱导的囊泡聚集和融合，Cer抑制水解活性。这些影响可以用两种脂质的不同物理特性来解释。 DAG 增加双层流动性并减少脂质的横向分离，从而增加酶活性和底物对酶的可及性。 Cer 具有相反的作用，主要是因为它倾向于将鞘磷脂（一种酶底物）隔离到刚性结构域中，而这些刚性结构域可能不太容易被酶接触。 (C) 2009 Elsevier B.V. 保留所有权利。
A phospholipase C/sphingomyelinase from Pseudomonas aeruginosa has been assayed on vesicles containing phosphatidylcholine, sphingomyelin, phosphatidylethanolamine and cholesterol at equimolar ratios. The enzyme activity modifies the bilayer chemical composition giving rise to diacylglycerol (DAG) and ceramide (Cer). Assays of enzyme activity, enzyme-induced aggregation and fusion have been performed. Ultrastructural evidence of vesicle fusion at various stages of the process is presented, based on cryo-EM observations. The two enzyme lipidic end-products, DAG and Cer, have opposite effects on the bilayer physical properties; the former abolishes lateral phase separation, while the latter generates a new gel phase [Sot et al., FEBS Lett. 582, 3230-3236 (2008)]. Addition of either DAG, or Cer, or both to the liposome mixture causes an increase in enzyme binding to the bilayers and a decrease in lag time of hydrolysis. These two lipids also have different effects on the enzyme activity, DAG enhancing enzyme-induced vesicle aggregation and fusion, Cer inhibiting the hydrolytic activity. These effects are explained in terms of the different physical properties of the two lipids. DAG increases bilayers fluidity and decreases lateral separation of lipids, thus increasing enzyme activity and substrate accessibility to the enzyme. Cer has the opposite effect mainly because of its tendency to sequester sphingomyelin, an enzyme substrate, into rigid domains, presumably less accessible to the enzyme. (C) 2009 Elsevier B.V. All rights reserved.