Health Effects Laboratory Division, National Institute for Occupational Safety and Health, Centers for Disease Control and Prevention, Morgantown, WV 26505, USA.
Anal Biochem. 2011 Jul 15;414(2):232-8. doi: 10.1016/j.ab.2011.03.035. Epub 2011 Mar 31.
Diisocyanates are highly reactive chemical compounds widely used in the manufacture of polyurethanes. Although diisocyanates have been identified as causative agents of allergic respiratory diseases, the specific mechanism by which these diseases occur is largely unknown. To better understand the chemical species produced when diisocyanates react with protein, tandem mass spectrometry was employed to unambiguously identify the binding sites of the industrially important isomers, 2,4- and 2,6-toluene diisocyanate, on human serum albumin at varying diisocyanate/protein ratios. The 2,4-isomer results in approximately 2-fold higher conjugation product ion abundances than does the 2,6-isomer, suggesting that the 2,4-isomer has a higher reactivity toward albumin. Both isomers preferentially react with the N-terminal amine of the protein and the ε-NH(2) of lysine. At a low (1:2) diisocyanate/protein ratio, five binding sites are identified, whereas at a high (40:1) ratio, near-stoichiometric conjugation is observed with a maximum of 37 binding sites identified. Binding sites observed at the lowest conjugation ratios are conserved at higher binding ratios, suggesting a subset of 5-10 preferential binding sites on albumin. Diisocyanate-protein conjugation results in a variety of reaction products, including intra- and intermolecular crosslinking, diisocyanate self-polymerization, and diisocyanate hydrolysis.
异氰酸酯是一种高度反应性的化学化合物，广泛应用于聚氨基甲酸酯的制造。虽然异氰酸酯已被确定为引起过敏性呼吸道疾病的原因，但这些疾病发生的确切机制在很大程度上尚不清楚。为了更好地了解异氰酸酯与蛋白质反应时产生的化学物质，串联质谱法被用来明确鉴定工业上重要的异构体 2,4-和 2,6-甲苯二异氰酸酯与人血清白蛋白在不同异氰酸酯/蛋白质比例下的结合部位。2,4-异构体的共轭产物离子丰度约比 2,6-异构体高 2 倍，这表明 2,4-异构体对白蛋白的反应性更高。两种异构体都优先与蛋白质的 N-末端胺和赖氨酸的 ε-NH(2)反应。在低（1:2）异氰酸酯/蛋白质比例下，鉴定出 5 个结合部位，而在高（40:1）比例下，观察到接近化学计量的共轭，鉴定出最多 37 个结合部位。在最低共轭比下观察到的结合部位在较高的结合比下保持不变，这表明白蛋白上有 5-10 个优先结合部位的亚群。异氰酸酯-蛋白质共轭会产生多种反应产物，包括内分子和分子间交联、异氰酸酯自聚合和异氰酸酯水解。
