Interdisciplinary Research Center HALOmem, Charles Tanford Protein Center, Institute for Biochemistry and Biotechnology, Martin Luther University Halle-Wittenberg, Halle, Germany.
Methods Mol Biol. 2021;2228:385-400. doi: 10.1007/978-1-0716-1024-4_26.
Cross-linking, in general, involves the covalent linkage of two amino acid residues of proteins or protein complexes in close proximity. Mass spectrometry and computational analysis are then applied to identify the formed linkage and deduce structural information such as distance restraints. Quantitative cross-linking coupled with mass spectrometry is well suited to study protein dynamics and conformations of protein complexes. The quantitative cross-linking workflow described here is based on the application of isotope labelled cross-linkers. Proteins or protein complexes present in different structural states are differentially cross-linked using a "light" and a "heavy" cross-linker. The intensity ratios of cross-links (i.e., light/heavy or heavy/light) indicate structural changes or interactions that are maintained in the different states. These structural insights lead to a better understanding of the function of the proteins or protein complexes investigated. The described workflow is applicable to a wide range of research questions including, for instance, protein dynamics or structural changes upon ligand binding.
交联，一般来说，涉及到蛋白质或蛋白质复合物中两个邻近的氨基酸残基的共价连接。然后应用质谱和计算分析来识别形成的键，并推断结构信息，如距离约束。定量交联与质谱联用非常适合研究蛋白质动力学和蛋白质复合物的构象。这里描述的定量交联工作流程基于同位素标记交联剂的应用。使用“轻”和“重”交联剂对处于不同结构状态的蛋白质或蛋白质复合物进行差异交联。交联的强度比（即轻/重或重/轻）表明在不同状态下保持的结构变化或相互作用。这些结构上的见解有助于更好地理解所研究的蛋白质或蛋白质复合物的功能。所描述的工作流程适用于广泛的研究问题，包括例如配体结合时的蛋白质动力学或结构变化。
