Department of Chemical Sciences, University of Naples Federico II, Naples, Italy.
Private Practice, Milton, MA, USA.
J Proteomics. 2021 Jan 16;231:104039. doi: 10.1016/j.jprot.2020.104039. Epub 2020 Nov 2.
Identification and characterization of ancient proteins still require technical developments towards non-invasiveness, sensitivity, versatility and ease of use of the analyses. We report that the enzyme functionalized films, described in Cicatiello et al. (2018), can be used efficiently on the surface of different objects ranging from fixative-coated paper to canvas to the coating on an albumen photograph, as well as the much harder surfaces of ivory objects and the proteinaceous binders in the decoration of a wooden Egyptian coffin. The mixture of digested peptides that are efficiently captured on the functionalized surface are also amenable to LC-MS/MS analysis, which is necessary to confidently identify chemical modifications induced upon degradation, in order to characterize the conservation state of proteins. Moreover, in a two-step procedure, we have combined the trypsin functionalized film with a PNGaseF functionalized film, which adds a deglycosylation pretreatment allowing improved detection of glycosylated proteins. SIGNIFICANCE: User friendly trypsin functionalized films were implemented to expand their potential as versatile, modular tools that can be widely exploited in the world of diagnosis of cultural heritage objects, ancient proteins, and palaeoproteomics: a procedure that could be carried out by conservators or archaeologists first on-site and later analysed with standard MS techniques.
鉴定和描述古代蛋白质仍然需要技术上的发展，以实现分析的非侵入性、敏感性、通用性和易用性。我们报告称，Cicatiello 等人（2018 年）所描述的酶功能化膜可有效地应用于从固定剂涂层纸到画布再到蛋白照片涂层的不同物体表面，以及象牙物体的更坚硬表面和木制埃及棺材装饰中的蛋白质结合剂。有效地被功能化表面捕获的消化肽混合物也适用于 LC-MS/MS 分析，这对于有信心地鉴定降解引起的化学修饰以确定蛋白质的保存状态是必要的。此外，我们在两步程序中，将胰蛋白酶功能化膜与 PNGaseF 功能化膜结合，这增加了一种糖基化预处理，允许改善对糖基化蛋白质的检测。意义：我们实施了用户友好的胰蛋白酶功能化膜，以扩大其作为通用、模块化工具的潜力，这些工具可广泛应用于文化遗产、古代蛋白质和古蛋白组学的诊断领域：这一程序可以由保护者或考古学家首先在现场进行，然后用标准的 MS 技术进行分析。
