Journal ArticleDOI
Proteome-wide profiling of protein assemblies by cross-linking mass spectrometry
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TLDR
An integrated workflow that robustly identifies cross-links from endogenous protein complexes in human cellular lysates is described, based on the application of mass spectrometry (MS)-cleavable cross-linkers, sequential collision-induced dissociation (CID)–tandem MS (MS/MS) and electron-transfer Dissociation (ETD)-MS/ MS acquisitions, and a dedicated search engine, XlinkX.Abstract:
A crosslinking-mass spectrometry strategy, including a new proteome database search engine called XlinkX, enables the identification of inter- and intra-protein cross-links in cell lysates on a proteome-wide scale. We describe an integrated workflow that robustly identifies cross-links from endogenous protein complexes in human cellular lysates. Our approach is based on the application of mass spectrometry (MS)-cleavable cross-linkers, sequential collision-induced dissociation (CID)–tandem MS (MS/MS) and electron-transfer dissociation (ETD)-MS/MS acquisitions, and a dedicated search engine, XlinkX, which allows rapid cross-link identification against a complete human proteome database. This approach allowed us to detect 2,426 unique cross-links at a 5% FDR (2,013 intraprotein and 413 interprotein cross-links) or 1,822 cross-links at a 1% FDR (1,622 intraprotein and 200 interprotein cross-links), indicating the detection of 326 or 134 protein-protein interactions at 5% FDR or 1% FDR, respectively, in HeLa cell lysates. We validated the confidence of our cross-linking results by using a target-decoy strategy and mapping the observed cross-link distances onto existing high-resolution structures. Our data provided new structural information about many protein assemblies and captured dynamic interactions of the ribosome in contact with different elongation factors.read more
Citations
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Journal ArticleDOI
Mass-spectrometric exploration of proteome structure and function
TL;DR: Powerful mass-spectrometry-based technologies now provide unprecedented insights into the composition, structure, function and control of the proteome, shedding light on complex biological processes and phenotypes.
Journal ArticleDOI
Mitochondrial proteins: from biogenesis to functional networks
TL;DR: How the mitochondrial protein import machinery functions as a key organizer of these protein networks, its involvement in the formation of membrane contact sites, and how defects in protein import can lead to disease are discussed.
Journal ArticleDOI
Crosslinking and Mass Spectrometry: An Integrated Technology to Understand the Structure and Function of Molecular Machines.
TL;DR: An overview of recent advances in XL-MS, the current state of the field, and a cursory outlook on future challenges is provided.
Journal ArticleDOI
A high-speed search engine pLink 2 with systematic evaluation for proteome-scale identification of cross-linked peptides
Zhen-Lin Chen,Jia-Ming Meng,Yong Cao,Yin Jili,Run-Qian Fang,Sheng-Bo Fan,Chao Liu,Wen-Feng Zeng,Yue-He Ding,Dan Tan,Long Wu,Wen-Jing Zhou,Hao Chi,Rui-Xiang Sun,Meng-Qiu Dong,Si-Min He +15 more
TL;DR: The pLink 2 as discussed by the authors is a search engine with higher speed and reliability for proteome-scale identification of cross-linked peptides, with a two-stage open search strategy facilitated by fragment indexing.
Journal ArticleDOI
Cross-Linking Mass Spectrometry: An Emerging Technology for Interactomics and Structural Biology.
Clinton Yu,Lan Huang +1 more
TL;DR: Strategies to Overcome Inherent Challenges in XLMS Studies and their Applications.
References
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Journal ArticleDOI
Minimal, encapsulated proteomic-sample processing applied to copy-number estimation in eukaryotic cells.
TL;DR: An in-StageTip method for performing sample processing, from cell lysis through elution of purified peptides, in a single, enclosed volume, and observes excellent quantitative reproducibility between replicates.
Journal ArticleDOI
Chemical cross-linking and mass spectrometry to map three-dimensional protein structures and protein-protein interactions.
TL;DR: The most popular cross-linking reagents for protein structure analysis are described and an overview of the different available strategies that employ chemical cross- linking and different mass spectrometric techniques are given.
Journal ArticleDOI
Identification of cross-linked peptides from complex samples
Bing Yang,Yan-Jie Wu,Ming Zhu,Sheng-Bo Fan,Jinzhong Lin,Kun Zhang,Shuang Li,Hao Chi,Yu-Xin Li,Hai-Feng Chen,Shukun Luo,Yue-He Ding,Le-Heng Wang,Zhiqi Hao,Li-Yun Xiu,She Chen,Keqiong Ye,Si-Min He,Meng-Qiu Dong +18 more
TL;DR: pLink as mentioned in this paper is a software for data analysis of cross-linked proteins coupled with mass-spectrometry analysis, which is compatible with multiple homo- or hetero-bifunctional cross-linkers.
Journal ArticleDOI
Structures of the human and Drosophila 80S ribosome
Andreas M. Anger,Jean Paul Armache,Otto Berninghausen,Michael Habeck,Marion Subklewe,Daniel N. Wilson,Roland Beckmann +6 more
TL;DR: In this paper, the structure of metazoan 80S ribosomes with the translation factor eEF2, E-site transfer RNA and Stm1-like proteins has been determined.
Journal ArticleDOI
The molecular sociology of the cell
TL;DR: Using emerging experimental techniques, such as mass spectrometry of complexes and single-particle cryo-electron microscopy, to complement traditional biochemical and biophysical methods will uncover the molecular architectures, and perhaps even atomic models, of many protein complexes.