Dafnis Ioannis, Metso Jari, Zannis Vassilis I, Jauhiainen Matti, Chroni Angeliki
Institute of Biosciences and Applications, National Center for Scientific Research "Demokritos" , Agia Paraskevi 15310, Athens, Greece.
Genomics and Biomarkers Unit, Biomedicum, National Institute for Health and Welfare , Helsinki 00290, Finland.
Biochemistry. 2015 Sep 29;54(38):5856-66. doi: 10.1021/acs.biochem.5b00681. Epub 2015 Sep 17.
Phospholipid transfer protein (PLTP), a main protein in lipid and lipoprotein metabolism, exists in high-activity (HA-PLTP) and low-activity (LA-PLTP) forms in human plasma. Proper phospholipid transfer activity of PLTP is modulated by interactions with various apolipoproteins (apo) including apoE. The domains of apoE involved in interactions with PLTP are not known. Here we analyzed the capacity of recombinant apoE isoforms and apoE4 mutants with progressive carboxyl-terminal deletions to bind to and activate HA-PLTP and LA-PLTP. Our analyses demonstrated that lipid-free apoE isoforms bind to both HA-PLTP and LA-PLTP, resulting in phospholipid transfer activation, with apoE3 inducing the highest PLTP activation. The isoform-specific differences in apoE/PLTP binding and PLTP activation were abolished following apoE lipidation. Lipid-free apoE4[Δ(260-299)], apoE4[Δ(230-299)], apoE4[Δ(203-299)], and apoE4[Δ(186-299)] activated HA-PLTP by 120-160% compared to full-length apoE4. Lipid-free apoE4[Δ(186-299)] also activated LA-PLTP by 85% compared to full-length apoE4. All lipidated truncated apoE4 forms displayed a similar effect on HA-PLTP and LA-PLTP activity as full-length apoE4. Strikingly, lipid-free or lipidated full-length apoE4 and apoE4[Δ(186-299)] demonstrated similar binding capacity to LA-PLTP and HA-PLTP. Biophysical studies showed that the carboxyl-terminal truncations of apoE4 resulted in small changes of the structural or thermodynamic properties of lipidated apoE4, that were much less pronounced compared to changes observed previously for lipid-free apoE4. Overall, our findings show an isoform-dependent binding to and activation of PLTP by lipid-free apoE. Furthermore, the domain of apoE4 required for PLTP activation resides within its amino-terminal 1-185 region. The apoE/PLTP interactions can be modulated by the conformation and lipidation state of apoE.
磷脂转运蛋白（PLTP）是脂质和脂蛋白代谢中的一种主要蛋白质，在人血浆中以高活性（HA-PLTP）和低活性（LA-PLTP）形式存在。PLTP适当的磷脂转运活性受其与包括载脂蛋白E（apoE）在内的各种载脂蛋白相互作用的调节。目前尚不清楚apoE中与PLTP相互作用的结构域。在此，我们分析了重组apoE异构体和羧基末端逐步缺失的apoE4突变体与HA-PLTP和LA-PLTP结合并激活它们的能力。我们的分析表明，无脂质的apoE异构体与HA-PLTP和LA-PLTP均能结合，从而激活磷脂转运，其中apoE3诱导的PLTP激活作用最强。apoE脂质化后，apoE/PLTP结合及PLTP激活的异构体特异性差异消失。与全长apoE4相比，无脂质的apoE4[Δ(260 - 299)]、apoE4[Δ(230 - 299)]、apoE4[Δ(203 - 299)]和apoE4[Δ(186 - 299)]可使HA-PLTP激活120% - 160%。与全长apoE4相比，无脂质的apoE4[Δ(186 - 299)]还可使LA-PLTP激活85%。所有脂质化的截短apoE4形式对HA-PLTP和LA-PLTP活性的影响与全长apoE4相似。令人惊讶的是，无脂质或脂质化的全长apoE4和apoE4[Δ(186 - 299)]对LA-PLTP和HA-PLTP表现出相似的结合能力。生物物理研究表明，apoE4的羧基末端截短导致脂质化apoE4的结构或热力学性质发生微小变化，与之前观察到的无脂质apoE4的变化相比，这种变化要小得多。总体而言，我们的研究结果表明无脂质的apoE与PLTP的结合及激活具有异构体依赖性。此外，激活PLTP所需的apoE4结构域位于其氨基末端的1 - 185区域。apoE/PLTP相互作用可受apoE的构象和脂质化状态调节。
