Department of Chemistry, Indian Institute of Science Education and Research Bhopal, Bhopal Bypass Road, Bhauri, Bhopal, 462 066, India.
Department of Biological Sciences, Indian Institute of Science Education and Research Bhopal, Bhopal Bypass Road, Bhauri, Bhopal, 462 066, India.
Angew Chem Int Ed Engl. 2020 Jun 22;59(26):10332-10336. doi: 10.1002/anie.202000062. Epub 2020 Apr 20.
The necessity for precision labeling of proteins emerged during the efforts to understand and regulate their structure and function. It demands selective attachment of tags such as affinity probes, fluorophores, and potent cytotoxins. Here, we report a method that enables single-site labeling of a high-frequency Lys residue in the native proteins. At first, the enabling reagent forms stabilized imines with multiple solvent-accessible Lys residues chemoselectively. These linchpins create the opportunity to regulate the position of a second Lys-selective electrophile connected by a spacer. Consequently, it enables the irreversible single-site labeling of a Lys residue independent of its place in the reactivity order. The user-friendly protocol involves a series of steps to deconvolute and address chemoselectivity, site-selectivity, and modularity. Also, it delivers ordered immobilization and analytically pure probe-tagged proteins. Besides, the methodology provides access to antibody-drug conjugate (ADC), which exhibits highly selective anti-proliferative activity towards HER-2 expressing SKBR-3 breast cancer cells.
在理解和调节蛋白质结构和功能的过程中，蛋白质的精确定量标记变得非常必要。这需要选择性地连接标签，如亲和探针、荧光团和有效的细胞毒素。在这里，我们报告了一种方法，该方法可以实现天然蛋白质中高频 Lys 残基的单点标记。首先，使能试剂与多个溶剂可及的 Lys 残基形成稳定的亚胺，具有化学选择性。这些连接点为通过间隔基连接的第二个 Lys 选择性亲电试剂的位置调节创造了机会。因此，它能够实现 Lys 残基的不可逆单点标记，而与它们在反应性顺序中的位置无关。用户友好的方案涉及一系列步骤来解析和解决化学选择性、位置选择性和模块性。此外，它还提供了有序的固定化和分析纯的探针标记蛋白质。此外，该方法学还提供了抗体药物偶联物 (ADC) 的途径，其对表达 HER-2 的 SKBR-3 乳腺癌细胞具有高度选择性的抗增殖活性。
