4种交联肽段富集方法

为了实现低丰度交联肽段的高效鉴定，以提供尽可能丰富的交联信息用于细胞内蛋白质复合物原位构象分析，应用可透膜型氨基反应活性，带有炔基可富集基团的交联剂BSP对293T细胞进行细胞水平原位化学交联，对交联细胞分别使用三种不同断裂方式的叠氮-生物素分子进行蛋白和肽段水平的点击化学反应，实现交联肽段的高效富集。上述数据分别进行了实际人源细胞复杂样品中交联肽段的富集测试，并采用液相色谱-高分辨质谱联用平台进行交联数据的鉴定。To achieve efficient identification of low-abundance cross-linked peptides and provide as comprehensive cross-linking information as possible for in situ conformational analysis of intracellular protein complexes, membrane-permeable, amine-reactive cross-linker BSP with alkynyl-based enrichment groups was used to perform in situ chemical cross-linking on 293T cells at the cellular level. The cross-linked cells were respectively subjected to click chemistry reactions using three types of azide-biotin molecules with distinct cleavage modes for protein and peptide-level enrichment, thereby realizing efficient enrichment of cross-linked peptides. The obtained datasets were separately tested for enrichment of cross-linked peptides in complex samples from actual human cells, and the cross-linking data were identified using a liquid chromatography-high resolution mass spectrometry (LC-HRMS) platform.

搜集汇总

背景与挑战

背景概述

该数据集介绍了使用BSP交联剂对293T细胞进行原位化学交联，并通过三种不同断裂方式的叠氮-生物素分子实现交联肽段高效富集的方法。数据集包含液相色谱-高分辨质谱联用平台鉴定的交联数据，旨在支持蛋白质复合物原位构象分析研究。