Macromolecular crowding in the Escherichia coli periplasm maintains α-synuclein disorder
Macromolecular crowding in the Escherichia coli periplasm maintains α-synuclein disorder
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DOI：
10.1016/j.jmb.2005.11.033
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发表时间：
2006-02-03
影响因子：
5.6
通讯作者：
Pielak, GJ
中科院分区：
文献类型：
--
作者：
McNulty, BC;Young, GB;Pielak, GJ
天然无序的蛋白质α-突触核蛋白是路易体的主要成分，路易体是帕金森病的细胞标志。大多数对该蛋白质的研究都是在稀溶液中进行的，但其生物学相关的作用是在细胞内拥挤的环境中进行的。我们通过将活体大肠杆菌中获得的 NMR 数据与体外 NMR 数据相结合，研究了大分子拥挤对 α-突触核蛋白的影响。大肠杆菌周质中拥挤的环境阻止了在 35 摄氏度的稀溶液中检测到的构象变化。这种变化与流体动力学半径的增加和 N 端 100 个氨基酸残基二级结构的形成有关。通过防止这种温度引起的构象变化，大肠杆菌周质中的拥挤稳定了无序的单体。我们在体外用 300 g/l 牛血清白蛋白拥挤 a-突触核蛋白时获得了相同的稳定性，表明单独拥挤足以稳定无序的单体蛋白质。两种与疾病相关的变体（A30P 和 A53T）在稀溶液和大肠杆菌周质中表现相同。这些数据揭示了根据具体情况处理大分子拥挤影响的重要性。此外，我们的工作表明，细胞内的α-突触核蛋白可能无法实现离散的结构化蛋白质构象，这表明大分子拥挤是帕金森病病因中一个可能的因素，这一点通常被忽视。 (c) 2005 Elsevier Ltd. 保留所有权利。
The natively disordered protein alpha-synuclein is the primary component of Lewy bodies, the cellular hallmark of Parkinson's disease. Most studies of this protein are performed in dilute solution, but its biologically relevant role is performed in the crowded environment inside cells. We addressed the effects of macromolecular crowding on alpha-synuclein by combining NMR data acquired in living Escherichia coli with in vitro NMR data. The crowded environment in the E. coli periplasm prevents a conformational change that is detected at 35 degrees C in dilute solution. This change is associated with an increase in hydrodynamic radius and the formation of secondary structure in the N-terminal 100 amino acid residues. By preventing this temperature-induced conformational change, crowding in the E. coli periplasm stabilizes the disordered monomer. We obtain the same stabilization in vitro upon crowding a-synuclein with 300 g/l of bovine serum albumin, indicating that crowding alone is sufficient to stabilize the disordered, monomeric protein. Two disease-associated variants (A30P and A53T) behave in the same way in both dilute solution and in the E. coli periplasm. These data reveal the importance of approaching the effects of macromolecular crowding on a case-by-case basis. Additionally, our work shows that discrete structured protein conformations may not be achieved by alpha-synuclein inside cells, implicating the commonly overlooked aspect of macromolecular crowding as a possible factor in the etiology of Parkinson's disease. (c) 2005 Elsevier Ltd. All rights reserved.