Table_13_A Robust and Universal Metaproteomics Workflow for Research Studies and Routine Diagnostics Within 24 h Using Phenol Extraction, FASP Digest, and the MetaProteomeAnalyzer.XLSX

The investigation of microbial proteins by mass spectrometry (metaproteomics) is a key technology for simultaneously assessing the taxonomic composition and the functionality of microbial communities in medical, environmental, and biotechnological applications. We present an improved metaproteomics workflow using an updated sample preparation and a new version of the MetaProteomeAnalyzer software for data analysis. High resolution by multidimensional separation (GeLC, MudPIT) was sacrificed to aim at fast analysis of a broad range of different samples in less than 24 h. The improved workflow generated at least two times as many protein identifications than our previous workflow, and a drastic increase of taxonomic and functional annotations. Improvements of all aspects of the workflow, particularly the speed, are first steps toward potential routine clinical diagnostics (i.e., fecal samples) and analysis of technical and environmental samples. The MetaProteomeAnalyzer is provided to the scientific community as a central remote server solution at www.mpa.ovgu.de.

借助质谱技术（mass spectrometry）开展微生物蛋白质组研究，即宏蛋白质组学（metaproteomics），是同时评估医学、环境及生物技术应用场景中微生物群落分类组成与功能特征的核心技术。本研究提出一套优化后的宏蛋白质组学分析流程，采用更新的样本制备方案与新版本的元蛋白质组分析器（MetaProteomeAnalyzer）软件开展数据分析。为实现在24小时内快速分析多种不同样本，我们舍弃了依托多维分离技术（GeLC、MudPIT）获取高分辨率数据的分析路径。经优化后的流程可实现的蛋白质鉴定数量较此前流程提升至少两倍，分类注释与功能注释的数量也大幅增长。该流程各环节的优化（尤其是分析速度的提升），为其在常规临床诊断（如粪便样本）、技术样本与环境样本分析中的潜在应用迈出了关键第一步。本研究将元蛋白质组分析器（MetaProteomeAnalyzer）作为面向科研界的中央远程服务器解决方案，部署于www.mpa.ovgu.de。