Elucidating the higher‐order structure of biopolymers by structural probing and mass spectrometry: MS3D

Abstract

Chemical probing represents a very versatile alternative for studying the structure and dynamics of substrates that are intractable by established high-resolution techniques. The implementation of MS-based strategies for the characterization of probing products has not only extended the range of applicability to virtually all types of biopolymers, but has also paved the way for the introduction of new reagents that would not have been viable with traditional analytical platforms. As the availability of probing data is steadily increasing on the wings of the development of dedicated interpretation aids, powerful computational approaches have been explored to enable the effective utilization of such information to generate valid molecular models. This combination of factors has contributed to making the possibility of obtaining actual 3D structures by MS-based technologies (MS3D) a reality. Although approaches for achieving structure determination of unknown substrates or assessing the dynamics of known structures may share similar reagents and development trajectories, they clearly involve distinctive experimental strategies, analytical concerns, and interpretation paradigms. This Perspective offers a commentary on methods aimed at obtaining distance constraints for the modeling of full-fledged structures, while highlighting common elements, salient distinctions, and complementary capabilities exhibited by methods employed in dynamics studies. We discuss critical factors to be addressed for completing effective structural determinations and expose possible pitfalls of chemical methods. We survey programs developed for facilitating the interpretation of experimental data and discuss possible computational strategies for translating sparse spatial constraints into all-atom models. Examples are provided to illustrate how the concerted application of very diverse probing techniques can lead to the solution of actual biological substrates.

StavroX—A Software for Analyzing Crosslinked Products in Protein Interaction Studies

“…C hemical crosslinking in combination with mass spectrometry (MS) has emerged as an alternative strategy to derive 3D structural information of proteins [1][2][3][4][5][6][7][8][9], which is reflected by the abbreviation "MS3D" to describe this fruitful combination of both techniques [3]. Chemical crosslinking relies on the introduction of a covalent bond between functional groups of amino acids within a protein (for gaining insight into the conformation of a protein) or between different interaction partners (for elucidating interfaces in protein complexes) by a chemical reagent.…”

“…Among the currently available software for analyzing crosslinked products are General Protein/Mass Analysis for Windows (GPMAW) [22], CoolToolBox, a major upgrade of the VIRTUALMSLAB software program [23], xQuest [24], X-Link Identifier [25], xComb [26], and MS-Bridge, which is part of Protein Prospector [27]. Summaries of currently available crosslinking software are found in [3] and [28].…”

Selective enrichment and identification of cross-linked peptides to study 3-D structures of protein complexes by mass spectrometry

“…These methods give rise to low resolution structures, and by modeling, different modules of the protein or protein complex can be placed within the structure. To validate and optimize the models, distance constraints within a protein or between proteins can be obtained by chemical cross-linking and mass spectrometry [1][2][3][4][5][6][7][8]. Also results from information-driven docking approaches can be experimentally validated with cross-linking [9].…”

Mapping protein structural changes by quantitative cross-linking

“…38 39 40 41 1. Introduction 42 Chemical cross-linking in combination with mass spectrometry 43 (XL-MS) is an established and powerful tool for the elucidation of 44 three-dimensional protein structures which also provides valuable 45 information about protein-protein and protein-nucleic acid inter- 46 actions [1][2][3][4][5]. Current cross-linking strategies do not significantly 47 differ from early experiments in which the analysis of products 48 was limited to photometric and electrophoretic methods [6].…”