Characterization of Androctonus mauretanicus Venom and In Vitro Screening of SARS-CoV-2 Entry Inhibitors Candidates

The project focuses on the characterization and exploration of the bioactive components of Androctonus mauritanicus scorpion venom, with an emphasis on proteomic profiling and peptide identification. The overarching goal is to generate a comprehensive dataset of venom peptides and proteins, which can serve as a resource for understanding venom composition, discovering novel bioactive molecules, and facilitating comparative analyses with other scorpion species. Specimens were collected from a high-risk region for scorpion envenomation and maintained under controlled laboratory conditions. Venom was extracted via low-voltage electrical stimulation, centrifuged to remove impurities, lyophilized, and stored at −80°C to preserve biological activity. The venom was then solubilized, desalted, and fractionated using solid-phase extraction and reverse-phase high-performance liquid chromatography, allowing for separation of complex protein mixtures into manageable fractions suitable for mass spectrometry analysis. Enzymatic digestion was performed using trypsin and Lys-C, following reduction and alkylation steps to break disulfide bonds and prevent their reformation. Peptide samples were concentrated and analyzed using high-resolution mass spectrometry, including both Quadrupole Time-of-Flight (Q-TOF) and Orbitrap Q-Exactive platforms, coupled with nano-flow liquid chromatography. Data-dependent acquisition strategies were employed to maximize peptide coverage, and advanced software tools were used to process raw spectra, identify peptide sequences, and annotate post-translational modifications. The resulting dataset provides high-confidence peptide identifications, including sequence information suitable for deposition in public proteomics repositories. By sharing these data through ProteomeXchange, the project aims to enhance reproducibility, enable comparative studies, and support further research into venom-derived bioactive compounds with potential therapeutic applications.

本项目聚焦于毛里塔尼亚杀人蝎（Androctonus mauritanicus）蝎毒的活性成分表征与探索，重点开展蛋白质组学谱分析与肽段鉴定工作。其总体目标为构建一套全面的蝎毒肽类与蛋白质数据集，该数据集可作为研究蝎毒组成、发现新型生物活性分子，并助力开展与其他蝎类物种比较分析的参考资源。实验所用蝎标本采集自蝎蛰伤高风险区域，并在标准化受控实验室环境中饲养。采用低压电刺激法提取蝎毒，经离心去除杂质后进行冷冻干燥，并置于−80℃低温环境中保存以维持其生物活性。随后将蝎毒进行溶解、脱盐，并通过固相萃取（solid-phase extraction）与反相高效液相色谱（reverse-phase high-performance liquid chromatography）进行分级分离，可将复杂的蛋白质混合物分离为适于质谱分析的可控级分。在完成还原与烷基化步骤以断裂二硫键并阻断其重新形成后，使用胰蛋白酶（trypsin）与Lys-C完成酶解消化。对肽段样品进行浓缩后，采用高分辨率质谱平台进行分析，包括四极杆飞行时间（Quadrupole Time-of-Flight, Q-TOF）与轨道阱Q-Exactive两种质谱平台，并联用纳流液相色谱系统。采用数据依赖性采集策略以最大化肽段覆盖度，并使用高级软件工具处理原始质谱谱图、鉴定肽段序列并注释翻译后修饰（post-translational modifications）。本数据集可提供高可信度的肽段鉴定结果，包含可提交至公共蛋白质组学数据库的序列信息。本项目计划通过ProteomeXchange（蛋白质组交换计划）共享这些数据，旨在提升研究可重复性、支持比较研究，并推动针对具有潜在治疗应用价值的蝎源活性化合物的后续研究。