Analysis of bilirubin-induced bovine serum albumin stability using protein denaturation and quantitative cross-linking mass spectrometry

We report the combination of protein-denaturation stability principles with quantitative cross-linking mass spectrometry using isobaric quantitative protein interaction reporter technologies. This method enables the evaluation of ligand-induced protein engagement through analysis of cross-link relative ratios across chemical denaturation. We found ligand-stabilized cross-linked lysine pairs in bovine serum albumin (BSA) and ligand bilirubin map to known binding sites Sudlow Site I and subdomain IB.

本研究将蛋白质变性稳定性原理与采用同量异位定量蛋白质相互作用报告物技术（isobaric quantitative protein interaction reporter technologies）的定量交联质谱（quantitative cross-linking mass spectrometry）相结合，并对该组合方法进行了报道。该方法可通过分析化学变性过程中的交联相对比率，实现对配体诱导的蛋白质结合状态的评估。本研究发现，牛血清白蛋白（BSA）与配体胆红素的结合体系中，经配体稳定的交联赖氨酸对均定位至已知结合位点萨德洛位点I（Sudlow Site I）与亚结构域IB。