Mass spectrometry locates local and allosteric conformational changes that occur on cofactor binding.
Mass spectrometry locates local and allosteric conformational changes that occur on cofactor binding.
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DOI：
10.1038/ncomms12163
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发表时间：
2016-07-15
影响因子：
16.6
通讯作者：
Barran PE
中科院分区：
文献类型：
--
作者：
Beveridge R;Migas LG;Payne KAP;Scrutton NS;Leys D;Barran PE
Fdc1 是一种脱羧酶，需要新型异戊二烯化 FMN 辅因子才能发挥活性。在这里，我们使用它作为示例系统来展示天然自上而下和自下而上质谱如何测量蛋白质与辅因子结合的结构效应。对于 Fdc1Ubix，辅因子赋予酶结构稳定性。 IM-MS 显示全息蛋白存在于四个密切相关的构象家族中，这些家族的 apo 形式不同；两个较小的家族在辅因子存在时人口较多，而在辅因子不存在时人口减少。这些发现得到 MD 模拟的支持，表明 apo 形式具有更开放的结构。 HDX-MS 揭示，虽然主要的结构变化发生在辅因子结合位点附近，但辅因子结合的重排在整个蛋白质中都很明显，主要归因于变构构象紧缩，与 IM-MS 数据一致。 脱羧酶 Fdc1 需要异戊二烯化黄素单核苷酸辅因子才能发挥活性。在这里，作者使用各种质谱技术来观察该蛋白质响应辅因子结合而发生的结构变化。
Fdc1 is a decarboxylase enzyme that requires the novel prenylated FMN cofactor for activity. Here, we use it as an exemplar system to show how native top-down and bottom-up mass spectrometry can measure the structural effect of cofactor binding by a protein. For Fdc1Ubix, the cofactor confers structural stability to the enzyme. IM–MS shows the holo protein to exist in four closely related conformational families, the populations of which differ in the apo form; the two smaller families are more populated in the presence of the cofactor and depopulated in its absence. These findings, supported by MD simulations, indicate a more open structure for the apo form. HDX-MS reveals that while the dominant structural changes occur proximal to the cofactor-binding site, rearrangements on cofactor binding are evident throughout the protein, predominantly attributable to allosteric conformational tightening, consistent with IM–MS data. The decarboxylate enzyme Fdc1 requires a prenylated flavin mononucleotide co-factor for activity. Here, the authors use a variety of mass spectrometric techniques to observe the structural changes in this protein in response to co-factor binding.