Photoredox-Catalyzed Decarboxylative C-Terminal Differentiation for Bulk- and Single-Molecule Proteomics.
Photoredox-Catalyzed Decarboxylative C-Terminal Differentiation for Bulk- and Single-Molecule Proteomics.
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DOI：
10.1021/acschembio.1c00631
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发表时间：
2021-11-19
影响因子：
4
通讯作者：
Anslyn, Eric, V
中科院分区：
文献类型：
--
作者：
Zhang, Le;Floyd, Brendan M.;Chilamari, Maheshwerreddy;Mapes, James;Swaminathan, Jagannath;Bloom, Steven;Marcotte, Edward M.;Anslyn, Eric, V
用于肽上的生物源官能团的选择性标记的方法正在被开发并用于当前和新兴的蛋白质组学技术（例如单分子荧光测序）的工作流程中。为了用任何一种方法实现成功的标记，需要肽片段含有标记化学所设计的官能团。在实践中，在每个肽片段中仅存在两个官能团，而不管蛋白质切割位点，即N-末端胺和C-末端羧酸。因此，开发全局标记技术需要特异性靶向肽的N-和/或C-末端。在这项工作中，我们展示了第一次成功应用光催化的C-末端脱羧烷基化肽质谱和单分子蛋白质测序，可以广泛应用于任何蛋白质组。我们证明，从牛血清白蛋白的酶促反应生成的复杂混合物中的肽，以及从酵母和人细胞提取物的蛋白质混合物，可以在其C-末端残基与迈克尔受体的位点特异性标记。使用两种不同的分析方法，我们的特点是C-末端标记效率大于50%的完整的蛋白质组和文件的倾向，各种C-末端氨基酸残基的脱羧标记，显示组氨酸和色氨酸是最不利的。最后，我们将联合收割机C-末端脱羧标记与正交羧酸标记技术串联起来，建立了一个新的荧光测序平台。
Methods for the selective labeling of biogenic functional groups on peptides are being developed and used in the workflow of both current and emerging proteomics technologies, such as single-molecule fluorosequencing. To achieve successful labeling with any one method requires that the peptide fragments contain the functional group for which labeling chemistry is designed. In practice, only two functional groups are present in every peptide fragment regardless of the protein cleavage site, namely, an N-terminal amine and a C-terminal carboxylic acid. Developing a global-labeling technology, therefore, requires one to specifically target the N- and/or C-terminus of peptides. In this work, we showcase the first successful application of photocatalyzed C-terminal decarboxylative alkylation for peptide mass spectrometry and single-molecule protein sequencing that can be broadly applied to any proteome. We demonstrate that peptides in complex mixtures generated from enzymatic digests from bovine serum albumin, as well as protein mixtures from yeast and human cell extracts, can be site-specifically labeled at their C-terminal residue with a Michael acceptor. Using two distinct analytical approaches, we characterize C-terminal labeling efficiencies of greater than 50% across complete proteomes and document the proclivity of various C-terminal amino-acid residues for decarboxylative labeling, showing histidine and tryptophan to be the most disfavored. Finally, we combine C-terminal decarboxylative labeling with an orthogonal carboxylic acid-labeling technology in tandem to establish a new platform for fluorosequencing.
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小肽通过光毒性催化的氧化C末端修饰。
影响因子：
8.4
作者：
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3
作者：
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