Quantitative peptidomics in mice - Effect of cocaine treatment
Quantitative peptidomics in mice - Effect of cocaine treatment
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DOI：
10.1385/jmn:28:3:265
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发表时间：
2006-01-01
影响因子：
3.1
通讯作者：
Fricker, LD
中科院分区：
文献类型：
--
作者：
Che, FY;Vathy, I;Fricker, LD
我们最近开发了一种定量肽组学方法，使用稳定同位素标记和质谱法来定量和鉴定大量肽。为了测试这种方法并筛选可卡因给药调节的肽，32 只 Cpe(fat/fat) 小鼠和 16 只野生型小鼠每天用盐水或 10 mg/kg 可卡因治疗两次，持续 5 天。从纹状体、下丘脑、海马和前额皮质中提取肽，并用含有九个氘或九个氢原子的三甲基丁酸铵的 N-羟基琥珀酰亚胺酯标记来自八只小鼠的组的提取物。将重标记和轻标记的肽池合并，在脱水胰蛋白酶亲和柱上纯化，并在与电喷雾电离四极杆飞行时间质谱仪耦合的反相柱上进行分析。根据可卡因与盐水峰的相对峰强度确定可卡因处理后肽水平的变化，并根据碰撞诱导解离光谱鉴定肽。在来自不同小鼠组的两次单独分析中，发现十种肽增加或减少。发现增加的肽对应于下丘脑中脑啡肽原、促甲状腺激素释放激素、加压素原、proSAAS、促分泌素 II、嗜铬素 B 和肽基甘氨酸-α-酰胺化单加氧酶的片段。前额皮质中相同的肽基-甘氨酸-α-酰胺化单加氧酶肽以及纹状体神经激肽 B 和两种未鉴定的肽均减少。在两次重复中，其他三十种肽均未受到可卡因处理的显着影响。总而言之，定量肽组学方法提供了一种筛选大量肽变化的有效方法。
We recently developed a quantitative peptidomics method using stable isotopic labels and mass spectrometry to both quantify and identify a large number of peptides. To test this approach and screen for peptides regulated by cocaine administration, 32 Cpe(fat/fat) mice and 16 wild-type mice were treated twice daily for 5 d either with saline or 10 mg/kg cocaine. Peptides were extracted from striatum, hypothalamus, hippocampus, and prefrontal cortex, and extracts from groups of eight mice were labeled with the N-hydroxysuccinimide ester of trimethylammonium butyrate containing either nine deuterium or nine hydrogen atoms. Pools of heavy- and light-labeled peptides were combined, purified on an anhydrotrypsin affinity column, and analyzed on a reverse-phase column coupled to an electrospray ionization quadrapole time-of-flight mass spectrometer. Changes in peptide levels upon cocaine treatment were determined from the relative peak intensities of the cocaine versus saline peaks, and peptides were identified from collision-induced dissociation spectra. Ten peptides were found to increase or decrease in each of two separate analyses from distinct groups of mice. Peptides found to increase corresponded to fragments of proenkephalin, prothyrotropin-releasing hormone, provasopressin, proSAAS, secretogranin II, chromogranin B, and peptidyl-glycine-alpha-amidating mono-oxygenase in the hypothalamus. The same peptidyl-glycine-alpha-amidating mono-oxygenase peptide decreased in the prefrontal cortex, along with striatal neurokinin B and two unidentified peptides. Thirty other peptides were not substantially affected by cocaine treatment in both replicates. Taken together, the quantitative peptidomics approach provides an efficient method to screen for changes in a large number of peptides.