Bound anions differentially stabilize multiprotein complexes in the absence of bulk solvent.
Bound anions differentially stabilize multiprotein complexes in the absence of bulk solvent.
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DOI：
10.1021/ja203527a
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发表时间：
2011-07-27
影响因子：
15
通讯作者：
Ruotolo, Brandon T.
中科院分区：
文献类型：
--
作者：
Han, Linjie;Hyung, Suk-Joon;Mayers, Jonathan J. S.;Ruotolo, Brandon T.
离子淌度分离与质谱的结合是一种新兴且强大的结构生物学工具，能够同时评估复杂混合物中大型蛋白质组装体的结构、拓扑、动力学和组成。离子淌度-质谱测量的一个组成部分是完整多蛋白复合物的电离以及它们从本体溶剂中的去除。在这个过程中，大部分蛋白质结构和组织可能被保留，给蛋白质施加了一个可能导致结构重排发生的外来环境。因此，必须确定一种通用方法来在没有大量溶剂的情况下稳定蛋白质结构。我们解决这个问题的方法涉及通过在解吸/电离之前在溶液中添加盐来保护蛋白质复合物结构。添加的盐的阴离子组分在溶液中或在电喷雾过程中与络合物结合，并且在气相中保持结合的那些往往具有高气相酸度。由此产生的反离子“壳”能够在激活时带走蛋白质复合离子的多余能量，并可导致气相蛋白质组装体整体结构的显着稳定。通过使用离子淌度-质谱法，我们观察了使用碰撞激活与十二种不同阴离子群结合的四种四聚蛋白复合物的解离和解折叠转变。这里提供的数据首次量化了大范围反离子对气相蛋白质结构的影响，并使我们能够在没有本体溶剂的情况下对作为结构稳定剂的反离子进行排序和分类。我们的测量表明，酒石酸根、柠檬酸根、氯离子和硝酸根阴离子是本次筛选中确定的气相蛋白质结构最强的稳定剂之一。与溶液中已知的霍夫迈斯特盐系列相比，由我们的数据确定的排名顺序有很大不同。虽然这是我们工作的预期结果，但由于水对阴离子和蛋白质溶剂化的影响减弱，我们的数据与溶液中预期的阴离子结合良好相关，并强调了水化层和阴离子-蛋白质结合效应对于溶液中霍夫迈斯特型稳定至关重要的事实。最后，我们提出了气相中反离子稳定蛋白质及其复合物的详细作用机制，这表明阴离子必须以高亲和力结合，但必须易于与蛋白质解离才能成为有效的稳定剂。针对较小蛋白质系统获得的阴离子解析数据使我们能够根据阴离子在没有大量溶剂的情况下稳定蛋白质和蛋白质复合物结构的能力将阴离子分为三类。
The combination of ion mobility separation with mass spectrometry is an emergent and powerful structural biology tool, capable of simultaneously assessing the structure, topology, dynamics and composition of large protein assemblies within complex mixtures. An integral part of the ion mobility-mass spectrometry measurement is the ionization of intact multiprotein complexes and their removal from bulk solvent. This process, during which a substantial portion of protein structure and organization is likely to be preserved, imposes a foreign environment on proteins that may cause structural rearrangements to occur. Thus, a general means must be identified to stabilize protein structures in the absence of bulk solvent. Our approach to this problem involves the protection of protein complex structure through the addition of salts in solution prior to desorption/ionization. Anionic components of the added salts bind to the complex either in solution or during the electrospray process, and those that remain bound in the gas phase tend to have high gas phase acidities. The resulting ‘shell’ of counter-ions is able to carry away excess energy from the protein complex ion upon activation and can result in significant structural stabilization of the gas-phase protein assembly overall. By using ion mobility-mass spectrometry, we observe both the dissociation and unfolding transitions for four tetrameric protein complexes bound to populations of twelve different anions using collisional activation. The data presented here quantifies, for the first time, the influence of a large range of counter-ions on gas-phase protein structure and allows us to rank and classify counter-ions as structure stabilizers in the absence of bulk solvent. Our measurements indicate that tartrate, citrate, chloride and nitrate anions are amongst the strongest stabilizers of gas-phase protein structure identified in this screen. The rank order determined by our data is substantially different when compared to the known Hofmeister salt series in solution. While this is an expected outcome of our work, due to the diminished influence of anion and protein solvation by water, our data correlates well to expected anion binding in solution and highlights the fact that both hydration layer and anion-protein binding effects are critical for Hofmeister-type stabilization in solution. Finally, we present a detailed mechanism of action for counter-ion stabilization of proteins and their complexes in the gas-phase, which indicates that anions must bind with high-affinity, but must dissociate readily from the protein in order to be an effective stabilizer. Anion-resolved data acquired for smaller protein systems allows us to classify anions into three categories based on their ability to stabilize protein and protein complex structure in the absence of bulk solvent.
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影响因子：
15
作者：
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通讯作者：
Robinson, Carol V.