Quantitative mass spectrometry measurements reveal stoichiometry of principal postsynaptic density proteins.
Quantitative mass spectrometry measurements reveal stoichiometry of principal postsynaptic density proteins.
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DOI：
10.1021/acs.jproteome.5b00109
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发表时间：
2015-06-05
影响因子：
4.4
通讯作者：
Dosemeci A
中科院分区：
文献类型：
--
作者：
Lowenthal MS;Markey SP;Dosemeci A
对大鼠大脑皮层突触后密度(PSD)组分进行了定量研究，最终目的是定义PSD复合体中蛋白质的平均拷贝数。利用同位素标记的多肽串联内标，开发了高特异性和选择性的同位素稀释质谱分析方法。PSD蛋白化学计量学的解释为PSD-95(SAP-90，DLG4)的摩尔比，随后，使用PSD-95的共识文献值估计拷贝数。PSD首次估计了几种蛋白质的平均拷贝数，包括AIDA-1、BRAGS和Densin的拷贝数。主要发现包括有证据表明PSD中AIDA-1的高拷贝数(144±30)-相当于GKAP总蛋白家族的拷贝数(150±27)-表明AIDA-1是PSD支架的一个元件。GluN1、GluN2A和GluN2B的NMDA受体亚基的平均拷贝数分别为66±18、27±9和45±15，共产生34±10个NMDA通道。AMPA通道及其辅助亚单位TARP的平均拷贝数估计分别为68±36和144±38，化学计量比约为1：2，支持大多数AMPA受体通过TARP亚单位锚定在PSD上的说法。这种对PSD蛋白质的可靠、定量的分析改进并扩展了PSD主要成分的列表，在解开PSD复杂的分子结构的持续努力中，分配了平均拷贝数。
Quantitative studies are presented of postsynaptic density (PSD) fractions from rat cerebral cortex with the ultimate goal of defining the average copy numbers of proteins in the PSD complex. Highly specific and selective isotope dilution mass spectrometry assays were developed using isotopically labeled polypeptide concatemer internal standards. Interpretation of PSD protein stoichiometry was achieved as a molar ratio with respect to PSD-95 (SAP-90, DLG4), and subsequently, copy numbers were estimated using a consensus literature value for PSD-95. Average copy numbers for several proteins at the PSD were estimated for the first time, including those for AIDA-1, BRAGs, and densin. Major findings include evidence for the high copy number of AIDA-1 in the PSD (144 ± 30)—equivalent to that of the total GKAP family of proteins (150 ± 27)—suggesting that AIDA-1 is an element of the PSD scaffold. The average copy numbers for NMDA receptor sub-units were estimated to be 66 ± 18, 27 ± 9, and 45 ± 15, respectively, for GluN1, GluN2A, and GluN2B, yielding a total of 34 ± 10 NMDA channels. Estimated average copy numbers for AMPA channels and their auxiliary sub-units TARPs were 68 ± 36 and 144 ± 38, respectively, with a stoichiometry of ~1:2, supporting the assertion that most AMPA receptors anchor to the PSD via TARP sub-units. This robust, quantitative analysis of PSD proteins improves upon and extends the list of major PSD components with assigned average copy numbers in the ongoing effort to unravel the complex molecular architecture of the PSD.
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影响因子：
3.7
作者：
Bordelon, JR;Smith, Y;Muly, EC
通讯作者：
Muly, EC
影响因子：
5.3
作者：
Dosemeci, A;Reese, TS;Cheng, JHT
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5.3
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作者：
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通讯作者：
Stein, Stephen E.
影响因子：
25
作者：
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通讯作者：
Ziff, Edward B.