Probing the Effects of Heterogeneous Oxidative Modifications on the Stability of Cytochrome c in Solution and in the Gas Phase.
Probing the Effects of Heterogeneous Oxidative Modifications on the Stability of Cytochrome c in Solution and in the Gas Phase.
复制标题
探讨多相氧化修饰对溶液和气相中细胞色素 c 稳定性的影响。
DOI：
--
复制
发表时间：
2020
影响因子：
3.2
通讯作者：
L. Konermann
中科院分区：
文献类型：
--
作者：
Victor C Yin;L. Konermann
活性氧的共价修饰可以调节蛋白质的功能和稳定性。溶液中的热展开实验是探测氧化引起的稳定性变化的标准工具。补充这样的解决方案的调查，电喷雾蛋白质离子的稳定性可以在气相中通过碰撞诱导展开（CIU）和离子迁移谱法进行评估。一个有待探讨的问题是，是否氧化诱导的稳定性改变在溶液中反映了气态蛋白质离子的CIU行为。在这里，我们解决这个问题，使用氯胺-T-氧化细胞色素c（CT-细胞色素c）作为模型系统。CT-cyt c包含经历MetO形成（+16 Da）和Lys羰基化（LysCH 2-NH 2-LysCHO，-1 Da）的各种蛋白形式。我们发现，溶液中的CT-cyt c是不稳定的，与未修饰的对照相比，熔解温度降低了约5 ℃。令人惊讶的是，CIU实验揭示了相反的趋势，即，CT-cyt c在气相中的稳定化。为了查明这种影响的来源，我们进行了蛋白质解析CIU的CT-细胞色素C馏分，已通过阳离子交换色谱分离。以这种方式，可以鉴定在残基80处的MetO形成为负责气相中稳定化的关键修饰。这种效应可能是由亚砜与蛋白质核心中的芳香残基新形成的接触引起的。总的来说，我们的研究结果表明，氧化修饰可以影响蛋白质在溶液中的稳定性和在气相中非常不同。
Covalent modifications by reactive oxygen species can modulate the function and stability of proteins. Thermal unfolding experiments in solution are a standard tool for probing oxidation-induced stability changes. Complementary to such solution investigations, the stability of electrosprayed protein ions can be assessed in the gas phase by collision-induced unfolding (CIU) and ion mobility spectrometry. A question that remains to be explored is whether oxidation-induced stability alterations in solution are mirrored by the CIU behavior of gaseous protein ions. Here we address this question using chloramine-T-oxidized cytochrome c (CT-cyt c) as a model system. CT-cyt c comprises various proteoforms that have undergone MetO formation (+16 Da) and Lys carbonylation (LysCH2-NH2 LysCHO, -1 Da). We found that CT-cyt c in solution was destabilized, with a ~5 C reduced melting temperature compared to unmodified controls. Surprisingly, CIU experiments revealed the opposite trend, i.e., a stabilization of CT-cyt c in the gas phase. To pinpoint the source of this effect, we performed proteoform-resolved CIU on CT-cyt c fractions that had been separated by cation exchange chromatography. In this way it was possible to identify MetO formation at residue 80 as the key modification responsible for stabilization in the gas phase. Possibly, this effect is caused by newly formed contacts of the sulfoxide with aromatic residues in the protein core. Overall, our results demonstrate that oxidative modifications can affect protein stability in solution and in the gas phase very differently.
登录
查看更多内容
硫醚和亚砜作为 N-乙酰微过氧化物酶 8 轴向配体的比较：对细胞色素 c 中蛋氨酸 80 氧化的影响。
影响因子：
4.6
作者：
Lushington,GeraldH;Cowley,AaronB;Silchenko,Svetlana;Lukat-Rodgers,GudrunS;Rodgers,KentonR;Benson,DavidR
通讯作者：
Benson,DavidR
影响因子：
2.9
作者：
Jayaraman, Shobini;Gantz, Donald L.;Gursky, Olga
通讯作者：
Gursky, Olga
甲硫氨酸氧化细胞色素的连接和反应性 c．
影响因子：
4.6
作者：
Zhong,Fangfang;Pletneva,EkaterinaV
通讯作者：
Pletneva,EkaterinaV
影响因子：
2.9
作者：
Belikova, NA;Vladimirov, YA;Kagan, VE
通讯作者：
Kagan, VE
DOI：
10.1016/j.bbabio.2015.06.006
发表时间：
2015-10
期刊：
Biochimica et biophysica acta
影响因子：
--
作者：
Birk AV;Chao WM;Liu S;Soong Y;Szeto HH
通讯作者：
Szeto HH