Correlated matrix-assisted laser desorption/ionization mass spectrometry and fluorescent imaging of photocleavable peptide-coded random bead-arrays
Correlated matrix-assisted laser desorption/ionization mass spectrometry and fluorescent imaging of photocleavable peptide-coded random bead-arrays
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DOI：
10.1002/rcm.6754
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发表时间：
2014-01-15
影响因子：
2
通讯作者：
Rothschild, Kenneth J.
中科院分区：
文献类型：
--
作者：
Lim, Mark J.;Liu, Ziying;Rothschild, Kenneth J.
理论依据快速进行蛋白质组学的全面筛选是后基因组时代的一个重要目标。相关基质辅助激光解吸电离质谱MALDI-MS和荧光成像的光裂解肽编码的随机珠阵列被评估为一个关键步骤，在一个新的方法，蛋白质组学筛选，结合了许多优点的MS与荧光为基础的微阵列。34 μ m直径的玻璃、琼脂糖或TentaGel（R）珠粒，使用光可裂解生物素或定制设计的光可裂解接头。将肽编码的珠库随机排列到具有45 μ m直径威尔斯的定制金包被微孔板中，并进行荧光和MALDI质谱成像（MALDI-MSI）。通过比较相同珠阵列的荧光和MALDI-MS图像，鉴定标记的珠并将其与其匹配的光可裂解质量标签相关联。通过MALDI-MSI检测，在光可切割肽编码的微珠阵列中的单个微珠暴露于蛋白激酶A（PKA）后，还检测到Kemptide肽的翻译后修饰。结论光可切割肽编码的微珠阵列的相关MALDI-MS和荧光成像可为进行全球蛋白质组学筛选提供基础。(c)2013作者质谱快速通讯（Rapid Communications in Mass Spectrometry），John Wiley & Sons，Ltd.
RATIONALERapidly performing global proteomic screens is an important goal in the post-genomic era. Correlated matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) and fluorescent imaging of photocleavable peptide-coded random bead-arrays was evaluated as a critical step in a new method for proteomic screening that combines many of the advantages of MS with fluorescence-based microarrays.METHODSSmall peptide-coded model bead libraries containing up to 20 different bead species were constructed by attaching peptides to 30-34 mu m diameter glass, agarose or TentaGel (R) beads using photocleavable biotin or a custom-designed photocleavable linker. The peptide-coded bead libraries were randomly arrayed into custom gold-coated micro-well plates with 45 mu m diameter wells and subjected to fluorescence and MALDI mass spectrometric imaging (MALDI-MSI).RESULTSPhotocleavable mass-tags from individual beads in these libraries were spatially localized as similar to 65 mu m spots using MALDI-MSI with high sensitivity and mass resolution. Fluorescently tagged beads were identified and correlated with their matching photocleavable mass-tags by comparing the fluorescence and MALDI-MS images of the same bead-array. Post-translational modification of the peptide Kemptide was also detected on individual beads in a photocleavable peptide-coded bead-array by MALDI-MSI alone, after exposure of the beads to protein kinase A (PKA).CONCLUSIONSCorrelated MALDI-MS and fluorescent imaging of photocleavable peptide-coded random bead-arrays can provide a basis for performing global proteomic screening. (c) 2013 The Authors. Rapid Communications in Mass Spectrometry published by John Wiley & Sons, Ltd.