P4-ATPase mechanism of phospholipid translocation

P4-ATP酶磷脂易位机制

基本信息

批准号：
8575204
负责人：
TODD R GRAHAM
金额：
$ 29.02万
依托单位：
VANDERBILT UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2013
资助国家：
美国
起止时间：
2013-09-01 至 2017-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8575204
关键词：
ATP Hydrolysis ATP phosphohydrolase ATP-Binding Cassette Transporters Active Biological Transport Amino Acids Apoptotic Binding Binding Sites Biochemical Blood Clot Blood coagulation Carrier Proteins Cations Cell Polarity Cell membrane Cells Characteristics Charge Chimera organism Cleaved cell Coupling Crosslinker Cysteine Cytokinesis Cytosol Data Defect Disease Environment Eukaryota Eukaryotic Cell Excision Face Family Glycosphingolipids Golgi Apparatus H(+)-K(+)-Exchanging ATPase Homology Modeling Human Immune Intrahepatic Cholestasis Ion Transport Learning Lecithin Lipids Male Infertility Malignant neoplasm of liver Mammalian Cell Mammals Maps Mediating Membrane Membrane Proteins Mental Retardation Modeling Molecular Genetics Mus Mutagenesis Mutation Na(+)-K(+)-Exchanging ATPase Non-Insulin-Dependent Diabetes Mellitus Obesity Pathway interactions Peripheral Phenylalanine Phosphatidylethanolamine Phosphatidylserines Phospholipids Positioning Attribute Progressive intrahepatic cholestasis Proteins Pump Resolution Role Saccharomycetales Series Side Signal Transduction Site Sphingolipids Structural Models Structure Subgroup Substrate Specificity System Testing Transmembrane Domain Vesicle Viral Water base crosslink extracellular hearing impairment human disease humoral immunity deficiency insight lipid transport member novel protein transport public health relevance screening

项目摘要

DESCRIPTION (provided by applicant): The defining feature of a cell is the incredibly thin sheet of membrane that demarcates the intracellular and extracellular milieus. This cell membrane, or plasma membrane, has a lipid component that provides a barrier to passage of polar or charged molecules, and protein component that allows passage of privileged molecules to make the membrane selectively permeable. Phospholipids are critical building blocks of the plasma membrane and these amphipathic molecules pack together side-by-side to form a two-layered sheet. The polar phospholipid headgroups in each leaflet face outward to interact with water and the hydrophobic fatty acyl chains face the interior of the bilayer structure. A remarkable characteristic of the eukaryotic cell membrane is that these two layers have a very different phospholipid composition, a phenomenon known as "membrane asymmetry". The inner leaflet facing the cytosol of mammalian cells is enriched in phosphatidylserine (PS) and phosphatidylethanolamine (PE) while the extracellular leaflet is enriched in sphingolipids, glycosphingolipids and phosphatidylcholine (PC). At least two classes of active-transport proteins are capable of moving phospholipids across a membrane bilayer to establish membrane asymmetry, including the type IV P-type ATPases (P4-ATPases) and members of the ATP-binding cassette (ABC) transporter families. Amazingly, even with decades of study and x-ray crystal structures of some ABC transporters, how lipids are transported remains a mystery. With ion- transporting P-type ATPases (type I, II and III), x-ray crystal structure have also been solved highlighting a structurally conserved substrate binding site in the center of the transmembrane domain (a canonical site). Whether P4-ATPases evolved the ability to recognize the much larger phospholipid substrate in this canonical site, or evolved a unique transport mechanism has been the subject of debate. Preliminary data supporting this proposal strongly suggests that the P4-ATPases are using a noncanonical transport pathway to flip their phospholipid substrate. We propose that phospholipid is being selected at both an "entry gate" near the extracellular leaflet and an "exit gate" near the cytosolic leaflet. Studies in this projet will test this two-gate hypothesis through mutational studies and better define the mechanism of phospholipid transport by the P4- ATPases. The information obtained will help us understand how human diseases arise from defects in membrane asymmetry or P4-ATPase deficiency.

描述（由申请人提供）：单元的定义特征是界限细胞内和细胞外环境的薄膜薄片。该细胞膜或质膜具有脂质成分，可为极性或带电分子的传递和蛋白质成分提供障碍，并允许通过特权分子传递使膜选择性地渗透性。磷脂是质膜的关键构件，这些两亲性分子并排包装在一起，形成两层薄板。每个小叶中的极性磷脂头组向外向外与水相互作用，疏水性脂肪酰基链面面向双层结构的内部。真核细胞膜的一个显着特征是，这两个层具有非常不同的磷脂组成，这是一种称为“膜不对称”的现象。面对哺乳动物细胞的胞质溶胶的内部小叶富含磷脂酰丝氨酸（PS）和磷脂酰乙醇胺（PE），而细胞外叶片则富含鞘脂，糖醇，糖磷脂脂和磷脂酰胆碱（PC）。至少有两类的活性传输蛋白能够在膜双层上移动磷脂，以建立膜不对称性，包括IV型P型ATPase（P4-ATPases）和ATP结合纸盒（ABC）转运蛋白的成员。令人惊讶的是，即使经过数十年的研究和某些ABC转运蛋白的X射线晶体结构，如何运输脂质仍然是一个谜。随着离子转运P型ATPase（I型，II和III），X射线晶体结构也已解决，突出了跨膜结构域（规范位点）中心的结构保守的底物结合位点。 P4-ATP酶是否进化了识别该规范部位中更大的磷脂底物的能力，还是发展出独特的运输机制的能力一直是争论的主题。支持该建议的初步数据强烈表明，P4-ATPase使用非规范的传输途径来翻转其磷脂底物。我们建议在细胞外叶片附近的“入口门”和胞质小叶附近的“出口门”中选择磷脂。该过程中的研究将通过突变研究检验这两个栅极假设，并更好地定义P4- ATPases磷脂转运的机制。获得的信息将有助于我们了解人类疾病是如何由于膜不对称或P4-ATPase缺乏症中缺陷引起的。