Extracting protein in its active form is critical for its functional characterization, and lipid removal is an essential step in the protein extraction process for further downstream applications. In the present study, we revisited the delipidation protocol and developed a rapid, solvent-free delipidation method using activated silica. The delipidated samples showed improved optical clarity and a significant reduction of endogenous lipids. The functional integrity of the lipases present in the delipidated sample was validated by in vitro enzyme assay using physiological substrate which includes neutral lipid as well as phospholipid. The accessibility of active site of the extracted enzymes was demonstrated by activity-based protein profiling (ABPP), a functional chemoproteomic approach. Detection of serine hydrolases using ABPP probe labeling was enhanced upon delipidation. Further, the total polyphenol content was significantly reduced, which helps to enhance the protein enrichment and small-molecule inhibitor screening by ABPP. Collectively, these results suggest that the present solvent-free delipidation approach is efficient and highly compatible with the functional characterization of the enzymes, particularly lipid hydrolases.

以活性形式提取蛋白质对其功能表征至关重要，而脂质去除是蛋白质提取过程中进一步下游应用的重要步骤。在本研究中，我们重新审视了脱脂方案，并开发了一种使用活性二氧化硅的快速、无溶剂脱脂方法。脱脂样品显示出改善的光学透明度和内源性脂质的显着减少。脱脂样品中存在的脂肪酶的功能完整性通过体外酶测定使用生理底物（包括中性脂质和磷脂）进行验证。通过基于活性的蛋白质分析 (ABPP)，一种功能性化学蛋白质组学方法，证明了提取酶活性位点的可及性。使用 ABPP 探针标记对丝氨酸水解酶的检测在脱脂后得到增强。此外，总多酚含量显着降低，这有助于增强 ABPP 对蛋白质的富集和小分子抑制剂的筛选。总的来说，这些结果表明，目前的无溶剂脱脂方法是有效的，并且与酶的功能特征，特别是脂质水解酶的功能特征高度兼容。