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As mass spectrometry (MS) offers readout of protein conformational changes at both the protein and peptide levels, MS\u2010based structural proteomics is making significant strides in the realms of structural and molecular biology, complementing traditional structural biology techniques. This review focuses on two powerful MS\u2010based techniques for peptide\u2010level readout, namely limited proteolysis\u2010mass spectrometry (LiP\u2010MS) and cross\u2010linking mass spectrometry (XL\u2010MS). First, we discuss the principles, features, and different workflows of these two methods. Subsequently, we delve into the bioinformatics strategies and software tools used for interpreting data associated with these protein conformation readouts and how the data can be integrated with other computational tools. Furthermore, we provide a comprehensive summary of the noteworthy applications of LiP\u2010MS and XL\u2010MS in diverse areas including neurodegenerative diseases, interactome studies, membrane proteins, and artificial intelligence\u2010based structural analysis. Finally, we discuss the factors that modulate protein conformational changes. We also highlight the remaining challenges in understanding the intricacies of protein conformational changes by LiP\u2010MS and XL\u2010MS technologies.<\/jats:p>","DOI":"10.1002\/mas.21908","type":"journal-article","created":{"date-parts":[[2024,9,20]],"date-time":"2024-09-20T01:50:47Z","timestamp":1726797047000},"page":"644-671","update-policy":"https:\/\/doi.org\/10.1002\/crossmark_policy","source":"Crossref","is-referenced-by-count":24,"title":["Mass Spectrometry Structural Proteomics Enabled by Limited Proteolysis and Cross\u2010Linking"],"prefix":"10.1002","volume":"45","author":[{"given":"Haiyan","family":"Lu","sequence":"first","affiliation":[{"name":"School of Pharmacy University of Wisconsin\u2010Madison Madison Wisconsin USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0009-0003-1605-3908","authenticated-orcid":false,"given":"Zexin","family":"Zhu","sequence":"additional","affiliation":[{"name":"School of Pharmacy University of 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