Use of top-down and bottom-up Fourier transform ion cyclotron resonance mass spectrometry for mapping calmodulin sites modified by platinum anticancer drugs.
Use of top-down and bottom-up Fourier transform ion cyclotron resonance mass spectrometry for mapping calmodulin sites modified by platinum anticancer drugs.
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DOI：
10.1021/ac202267g
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发表时间：
2011-12-15
影响因子：
7.4
通讯作者：
O'Connor, Peter B.
中科院分区：
文献类型：
--
作者：
Li, Huilin;Lin, Tzu-Yung;Van Orden, Steve L.;Zhao, Yao;Barrow, Mark P.;Pizarro, Ana M.;Qi, Yulin;Sadler, Peter J.;O'Connor, Peter B.
钙调素（CaM）是一种高度保守的、普遍存在的钙结合蛋白;它结合并调节许多不同的蛋白质靶点，从而起到钙传感器和信号转导器的作用。CaM含有9个甲硫氨酸（Met）、1个组氨酸（His）、17个天冬氨酸（Asp）和23个谷氨酰胺酸（Glu）残基，所有这些残基都可能与铂化合物发生反应;因此，CaM序列的三分之一是铂抗癌药物可能的结合靶点，这对识别特定的铂修饰位点提出了重大挑战。在这里，自上而下的电子捕获解离（ECD）被用来阐明过渡金属铂（II）的修改网站。采用自上而下和自下而上相结合的质谱方法，鉴定了单核配合物顺铂和[Pt（dien）Cl]Cl，双核配合物[{cis-PtCl 2（NH 3）}2（μ-NH 2（CH 2）4 NH 2）]在CaM上的10个特异性结合位点。顺铂修饰的CaM的高分辨率MS显示，顺铂主要靶向在溶液中的低摩尔比的顺铂-CaM（2：1），通过交联Met残基的Met残基。在顺铂：CaM（8：1）的高摩尔比下，多达10个铂（II）与Met、Asp和Glu残基结合。[{cis-PtCl 2（NH3）}2（μ-NH 2（CH 2）4 NH 2）]与CaM形成单核加合物。两个铂中心之间的链烷二胺连接基由于反式不稳定效应而解离。[Pt（dien）Cl]Cl与CaM形成{Pt（dien）}2+加合物，其优先结合位点被鉴定为Met 51，Met 71，Met 72，His 107，Met 109，Met 124，Met 144，Met 145，Glu 45或Glu 47，以及Asp 122或Glu 123。这些复合物与CaM的结合，特别是当结合涉及所有四种原始配体的丢失时，在很大程度上是不可逆的，这可能导致它们无法到达靶DNA或在化疗期间造成不必要的副作用。此外，顺铂与CaM的交联可能导致CaM或CaM-Ca 2+复合物的生物学功能丧失，这是由于限制了CaM或CaM-Ca 2+复合物识别靶蛋白的灵活性或阻断了靶蛋白与CaM的结合区域。
Calmodulin (CaM) is a highly conserved, ubiquitous, calcium-binding protein; it binds to and regulates many different protein targets, thereby functioning as a calcium sensor and signal transducer. CaM contains 9 methionine (Met), 1 histidine (His), 17 aspartic acid (Asp), and 23 glutamine acid (Glu) residues, all of which can potentially react with platinum compounds; thus, one third of the CaM sequence is a possible binding target of platinum anticancer drugs, which represents a major challenge for identification of specific platinum modification sites. Here, top-down electron capture dissociation (ECD) was used to elucidate the transition metal-platinum(II) modification sites. By using a combination of top-down and bottom-up mass spectrometric (MS) approaches, ten specific binding sites for mononuclear complexes, cisplatin and [Pt(dien)Cl]Cl, dinuclear complex [{cis-PtCl2(NH3)}2(μ-NH2(CH2)4NH2)] on CaM were identified. High resolution MS of cisplatin-modified CaM revealed that cisplatin mainly targets Met residues in solution at low molar ratios of cisplatin-CaM (2:1), by cross-linking Met residues. At a high molar ratio of cisplatin:CaM (8:1), up to 10 platinum(II) bind to Met, Asp, and Glu residues. [{cis-PtCl2(NH3)}2(μ-NH2(CH2)4NH2)] forms mononuclear adducts with CaM. The alkanediamine linker between the two platinum centres dissociates due to a trans-labilization effect. [Pt(dien)Cl]Cl forms {Pt(dien)}2+ adducts with CaM, and the preferential binding sites were identified as Met51, Met71, Met72, His107, Met109, Met124, Met144, Met145, Glu45 or Glu47, and Asp122 or Glu123. The binding of these complexes to CaM, particularly when binding involves loss of all four original ligands, is largely irreversible which could result in their failure to reach the target DNA or be responsible for unwanted side-effects during chemotherapy. Additionally, the cross-linking of cisplatin to CaM might lead to the loss of the biological function of CaM or CaM-Ca2+ due to limiting the flexibility of the CaM or CaM-Ca2+ complex to recognize target proteins or blocking the binding region of target proteins to CaM.
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影响因子：
4.6
作者：
Hartinger, Christian G.;Tsybin, Yury O.;Dyson, Paul J.
通讯作者：
Dyson, Paul J.
DOI：
10.1002/oms.1210260527
发表时间：
1991-05-01
期刊：
ORGANIC MASS SPECTROMETRY
影响因子：
--
作者：
CARAVATTI, P;ALLEMANN, M
通讯作者：
ALLEMANN, M
影响因子：
15
作者：
Kelleher, NL;Lin, HY;McLafferty, FW
通讯作者：
McLafferty, FW
影响因子：
7.3
作者：
Knipp, Markus;Karotki, Andrei V.;Vasak, Milan
通讯作者：
Vasak, Milan
影响因子：
2
作者：
Feketeova, Linda;Ryzhov, Victor;O'Hair, Richard A. J.
通讯作者：
O'Hair, Richard A. J.