Native Mass Spectrometry Analysis of Oligomerization States of Fluorescence Recovery Protein and Orange Carotenoid Protein: Two Proteins Involved in the Cyanobacterial Photoprotection Cycle
Native Mass Spectrometry Analysis of Oligomerization States of Fluorescence Recovery Protein and Orange Carotenoid Protein: Two Proteins Involved in the Cyanobacterial Photoprotection Cycle
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DOI：
10.1021/acs.biochem.6b01094
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发表时间：
2017-01-10
期刊：
影响因子：
2.9
通讯作者：
Blankenship, Robert E.
中科院分区：
文献类型：
--
作者：
Lu, Yue;Liu, Haijun;Blankenship, Robert E.
橙子类胡萝卜素蛋白（OCP）和荧光恢复蛋白（FRP）存在于许多蓝藻中，并以拮抗的方式作为光强度的函数调节基本的光保护循环。我们的特点是寡聚化状态的OCP和FRP通过使用本地质谱，一种技术，具有研究天然蛋白质在广泛的蛋白质浓度和分子量的能力。我们发现，二聚FRP是蛋白质浓度范围从3至180 μ M的主要状态，高阶低聚物逐渐形成在蛋白质浓度高于此范围。然而，OCP表现出显著不同的低聚行为。在低蛋白浓度下，单体OCP（mOCP）占主导地位，可观察到二聚体OCP（dOCP）。然而，dOCP与mOCP的比率随着蛋白质浓度成比例地增加。在蛋白质浓度超过10 μ M时形成更高级的OCP寡聚体。此外，本机质谱加上离子迁移率使我们能够测量蛋白质碰撞截面和询问不同的FRP和OCP寡聚体的展开。我们发现，单体FRP表现出一个阶段的展开过程，这可能与其C-末端弯曲的晶体结构。对FRP和OCP的结构域组成进行了比较和讨论。
The orange carotenoid protein (OCP) and fluorescence recovery protein (FRP) are present in many cyanobacteria and regulate an essential photoprotection cycle in an antagonistic manner as a function of light intensity. We characterized the oligomerization states of OCP and FRP by using native mass spectrometry, a technique that has the capability of studying native proteins under a wide range of protein concentrations and molecular masses. We found that dimeric FRP is the predominant state at protein concentrations ranging from 3 to 180 mu M and that higher-order oligomers gradually form at protein concentrations above this range. The OCP, however, demonstrates significantly different oligomerization behavior. Monomeric OCP (mOCP) dominates at low protein concentrations, with an observable population of dimeric OCP (dOCP). The ratio of dOCP to mOCP, however, increases proportionally with protein concentration. Higher-order OCP oligomers form at protein concentrations beyond 10 mu M. Additionally, native mass spectrometry coupled with ion mobility allowed us to measure protein collisional cross sections and interrogate the unfolding of different FRP and OCP oligomers. We found that monomeric FRP exhibits a one-stage unfolding process, which could be correlated with its C-terminal bent crystal structure. The structural domain compositions of FRP and OCP are compared and discussed.