Matrix-assisted nanoelectrospray mass spectrometry for soft ionization of metal(i)–protein complexes

The Analyst ◽  
2020 ◽  
Vol 145 (5) ◽  
pp. 1646-1656
Author(s):  
Jin Li ◽  
Yajun Zheng ◽  
Jia Zhao ◽  
Daniel E. Austin ◽  
Zhiping Zhang
Keyword(s):  
Mass Spectrometry ◽  
Metal Ions ◽  
Protein Interactions ◽  
Protein Complexes ◽  
Biological Processes ◽  
Soft Ionization

Metal ions play significant roles in biological processes, and investigation of metal–protein interactions provides a basis to understand the functions of metal ions in such systems.

scholarly journals RNA-Centric Approaches to Profile the RNA–Protein Interaction Landscape on Selected RNAs

2021 ◽  
Vol 7 (1) ◽  
pp. 11 ◽  
Author(s):  
André P. Gerber
Keyword(s):  
Mass Spectrometry ◽  
Protein Interactions ◽  
Regulatory Networks ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Protein Complexes ◽  
Cell Protein ◽  
Transcriptional Regulatory Networks ◽  
Technological Advances

RNA–protein interactions frame post-transcriptional regulatory networks and modulate transcription and epigenetics. While the technological advances in RNA sequencing have significantly expanded the repertoire of RNAs, recently developed biochemical approaches combined with sensitive mass-spectrometry have revealed hundreds of previously unrecognized and potentially novel RNA-binding proteins. Nevertheless, a major challenge remains to understand how the thousands of RNA molecules and their interacting proteins assemble and control the fate of each individual RNA in a cell. Here, I review recent methodological advances to approach this problem through systematic identification of proteins that interact with particular RNAs in living cells. Thereby, a specific focus is given to in vivo approaches that involve crosslinking of RNA–protein interactions through ultraviolet irradiation or treatment of cells with chemicals, followed by capture of the RNA under study with antisense-oligonucleotides and identification of bound proteins with mass-spectrometry. Several recent studies defining interactomes of long non-coding RNAs, viral RNAs, as well as mRNAs are highlighted, and short reference is given to recent in-cell protein labeling techniques. These recent experimental improvements could open the door for broader applications and to study the remodeling of RNA–protein complexes upon different environmental cues and in disease.

scholarly journals Mass spectrometry-based cross-linking study shows that the Psb28 protein binds to cytochrome b559 in Photosystem II

2017 ◽  
Vol 114 (9) ◽  
pp. 2224-2229 ◽  
Author(s):  
Daniel A. Weisz ◽  
Haijun Liu ◽  
Hao Zhang ◽  
Sundarapandian Thangapandian ◽  
Emad Tajkhorshid ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Photosystem Ii ◽  
Protein Interactions ◽  
Protein Complexes ◽  
Protein Docking ◽  
Cross Linking ◽  
Protein Protein Interactions ◽  
Cytochrome B559 ◽  
Reaction Center Complex ◽  
Assembly Intermediate

Photosystem II (PSII), a large pigment protein complex, undergoes rapid turnover under natural conditions. During assembly of PSII, oxidative damage to vulnerable assembly intermediate complexes must be prevented. Psb28, the only cytoplasmic extrinsic protein in PSII, protects the RC47 assembly intermediate of PSII and assists its efficient conversion into functional PSII. Its role is particularly important under stress conditions when PSII damage occurs frequently. Psb28 is not found, however, in any PSII crystal structure, and its structural location has remained unknown. In this study, we used chemical cross-linking combined with mass spectrometry to capture the transient interaction of Psb28 with PSII. We detected three cross-links between Psb28 and the α- and β-subunits of cytochrome b559, an essential component of the PSII reaction-center complex. These distance restraints enable us to position Psb28 on the cytosolic surface of PSII directly above cytochrome b559, in close proximity to the QB site. Protein–protein docking results also support Psb28 binding in this region. Determination of the Psb28 binding site and other biochemical evidence allow us to propose a mechanism by which Psb28 exerts its protective effect on the RC47 intermediate. This study also shows that isotope-encoded cross-linking with the “mass tags” selection criteria allows confident identification of more cross-linked peptides in PSII than has been previously reported. This approach thus holds promise to identify other transient protein–protein interactions in membrane protein complexes.

scholarly journals Mass spectrometry-based methods for analysing the mitochondrial interactome in mammalian cells

2019 ◽  
Vol 167 (3) ◽  
pp. 225-231 ◽  
Author(s):  
Takumi Koshiba ◽  
Hidetaka Kosako
Keyword(s):  
Mass Spectrometry ◽  
Protein Interactions ◽  
Mammalian Cells ◽  
Protein Complexes ◽  
Living Cells ◽  
Protein Protein Interactions ◽  
Cellular Functions ◽  
Protein Protein Interaction ◽  
Membrane Protein Complexes ◽  
Protein Protein Interaction Networks

Abstract Protein–protein interactions are essential biologic processes that occur at inter- and intracellular levels. To gain insight into the various complex cellular functions of these interactions, it is necessary to assess them under physiologic conditions. Recent advances in various proteomic technologies allow to investigate protein–protein interaction networks in living cells. The combination of proximity-dependent labelling and chemical cross-linking will greatly enhance our understanding of multi-protein complexes that are difficult to prepare, such as organelle-bound membrane proteins. In this review, we describe our current understanding of mass spectrometry-based proteomics mapping methods for elucidating organelle-bound membrane protein complexes in living cells, with a focus on protein–protein interactions in mitochondrial subcellular compartments.

scholarly journals PPI-MASS: An Interactive Web Server to Identify Protein-Protein Interactions From Mass Spectrometry-Based Proteomics Data

2021 ◽  
Vol 8 ◽  
Author(s):  
Mariela González-Avendaño ◽  
Simón Zúñiga-Almonacid ◽  
Ian Silva ◽  
Boris Lavanderos ◽  
Felipe Robinson ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Protein Interactions ◽  
Protein Complexes ◽  
Web Server ◽  
Target Protein ◽  
Protein Protein Interactions ◽  
Proteomics Data ◽  
Functional Protein ◽  
Web Platform ◽  
The Web

Mass spectrometry-based proteomics methods are widely used to identify and quantify protein complexes involved in diverse biological processes. Specifically, tandem mass spectrometry methods represent an accurate and sensitive strategy for identifying protein-protein interactions. However, most of these approaches provide only lists of peptide fragments associated with a target protein, without performing further analyses to discriminate physical or functional protein-protein interactions. Here, we present the PPI-MASS web server, which provides an interactive analytics platform to identify protein-protein interactions with pharmacological potential by filtering a large protein set according to different biological features. Starting from a list of proteins detected by MS-based methods, PPI-MASS integrates an automatized pipeline to obtain information of each protein from freely accessible databases. The collected data include protein sequence, functional and structural properties, associated pathologies and drugs, as well as location and expression in human tissues. Based on this information, users can manipulate different filters in the web platform to identify candidate proteins to establish physical contacts with a target protein. Thus, our server offers a simple but powerful tool to detect novel protein-protein interactions, avoiding tedious and time-consuming data postprocessing. To test the web server, we employed the interactome of the TRPM4 and TMPRSS11a proteins as a use case. From these data, protein-protein interactions were identified, which have been validated through biochemical and bioinformatic studies. Accordingly, our web platform provides a comprehensive and complementary tool for identifying protein-protein complexes assisting the future design of associated therapies.

scholarly journals A synthesis of over 9,000 mass spectrometry experiments reveals the core set of human protein complexes

2016 ◽  
Author(s):  
Kevin Drew ◽  
Chanjae Lee ◽  
Ryan L. Huizar ◽  
Fan Tu ◽  
Blake Borgeson ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Protein Interactions ◽  
Protein Complexes ◽  
Genetic Diseases ◽  
Intraflagellar Transport ◽  
Human Protein ◽  
Disease Genes ◽  
Cellular Functions ◽  
The Core

AbstractMacromolecular protein complexes carry out many of the essential functions of cells, and many genetic diseases arise from disrupting the functions of such complexes. Currently there is great interest in defining the complete set of human protein complexes, but recent published maps lack comprehensive coverage. Here, through the synthesis of over 9,000 published mass spectrometry experiments, we present hu.MAP, the most comprehensive and accurate human protein complex map to date, containing >4,600 total complexes, >7,700 proteins and >56,000 unique interactions, including thousands of confident protein interactions not identified by the original publications. hu.MAP accurately recapitulates known complexes withheld from the learning procedure, which was optimized with the aid of a new quantitative metric (k-cliques) for comparing sets of sets. The vast majority of complexes in our map are significantly enriched with literature annotations and the map overall shows improved coverage of many disease-associated proteins, as we describe in detail for ciliopathies. Using hu.MAP, we predicted and experimentally validated candidate ciliopathy disease genes in vivo in a model vertebrate, discovering CCDC138, WDR90, and KIAA1328 to be new cilia basal body/centriolar satellite proteins, and identifying ANKRD55 as a novel member of the intraflagellar transport machinery. By offering significant improvements to the accuracy and coverage of human protein complexes, hu.MAP (http://proteincomplexes.org) serves as a valuable resource for better understanding the core cellular functions of human proteins and helping to determine mechanistic foundations of human disease.

scholarly journals Reliable identification of protein-protein interactions by crosslinking mass spectrometry

2020 ◽  
Author(s):  
Swantje Lenz ◽  
Ludwig R. Sinn ◽  
Francis J. O’Reilly ◽  
Lutz Fischer ◽  
Fritz Wegner ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Protein Interactions ◽  
Large Scale ◽  
Protein Complexes ◽  
Estimation Procedure ◽  
Scale Analysis ◽  
Protein Protein Interactions ◽  
False Discovery ◽  
Large Scale Analysis ◽  
False Discovery Rate Estimation

Crosslinking mass spectrometry is widening its scope from structural analyzes of purified multi-protein complexes towards systems-wide analyzes of protein-protein interactions. Assessing the error in these large datasets is currently a challenge. Using a controlled large-scale analysis of Escherichia coli cell lysate, we demonstrate a reliable false-discovery rate estimation procedure for protein-protein interactions identified by crosslinking mass spectrometry.

scholarly journals Stitching the synapse: Cross-linking mass spectrometry into resolving synaptic protein interactions

2020 ◽  
Vol 6 (8) ◽  
pp. eaax5783 ◽  
Author(s):  
M. A. Gonzalez-Lozano ◽  
F. Koopmans ◽  
P. F. Sullivan ◽  
J. Protze ◽  
G. Krause ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Protein Interactions ◽  
Protein Complexes ◽  
Conformational Dynamics ◽  
Synaptic Proteins ◽  
Cross Linking ◽  
Synaptic Protein ◽  
Protein Partners ◽  
Associated Proteins ◽  
Model Protein

Synaptic transmission is the predominant form of communication in the brain. It requires functionally specialized molecular machineries constituted by thousands of interacting synaptic proteins. Here, we made use of recent advances in cross-linking mass spectrometry (XL-MS) in combination with biochemical and computational approaches to reveal the architecture and assembly of synaptic protein complexes from mouse brain hippocampus and cerebellum. We obtained 11,999 unique lysine-lysine cross-links, comprising connections within and between 2362 proteins. This extensive collection was the basis to identify novel protein partners, to model protein conformational dynamics, and to delineate within and between protein interactions of main synaptic constituents, such as Camk2, the AMPA-type glutamate receptor, and associated proteins. Using XL-MS, we generated a protein interaction resource that we made easily accessible via a web-based platform (http://xlink.cncr.nl) to provide new entries into exploration of all protein interactions identified.

scholarly journals Optimization of Formaldehyde Cross-Linking for Protein Interaction Analysis of Non-Tagged Integrinβ1

2010 ◽  
Vol 2010 ◽  
pp. 1-13 ◽  
Author(s):  
Cordula Klockenbusch ◽  
Juergen Kast
Keyword(s):  
Mass Spectrometry ◽  
Protein Interactions ◽  
Interaction Analysis ◽  
Protein Complexes ◽  
Human Platelets ◽  
Jurkat Cells ◽  
Mass Spectrometry Analysis ◽  
Cross Linking ◽  
High Molecular Weight Complex ◽  
Spectrometry Analysis

Formaldehyde cross-linking of protein complexes combined with immunoprecipitation and mass spectrometry analysis is a promising technique for analysing protein-protein interactions, including those of transient nature. Here we used integrinβ1 as a model to describe the application of formaldehyde cross-linking in detail, particularly focusing on the optimal parameters for cross-linking, the detection of formaldehyde cross-linked complexes, the utility of antibodies, and the identification of binding partners. Integrinβ1 was found in a high molecular weight complex after formaldehyde cross-linking. Eight different anti-integrinβ1 antibodies were used for pull-down experiments and no loss in precipitation efficiency after cross-linking was observed. However, two of the antibodies could not precipitate the complex, probably due to hidden epitopes. Formaldehyde cross-linked complexes, precipitated from Jurkat cells or human platelets and analyzed by mass spectrometry, were found to be composed of integrinβ1,α4 andα6 orβ1,α6,α2, andα5, respectively.

scholarly journals Synthesis of two new enrichable and MS-cleavable cross-linkers to define protein–protein interactions by mass spectrometry

2015 ◽  
Vol 13 (17) ◽  
pp. 5030-5037 ◽  
Author(s):  
Anthony M. Burke ◽  
Wynne Kandur ◽  
Eric J. Novitsky ◽  
Robyn M. Kaake ◽  
Clinton Yu ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Protein Interactions ◽  
Structural Information ◽  
Protein Complexes ◽  
Cross Linking ◽  
Protein Protein Interactions ◽  
The Cross

The cross-linking Mass Spectrometry (XL-MS) technique extracts structural information from protein complexes without requiring highly purified samples, crystallinity, or large amounts of material.

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