A protocol for studying structural dynamics of proteins by quantitative crosslinking mass spectrometry and data-independent acquisition

Müller, Friedrich; Rappsilber, Juri

doi:10.1016/j.jprot.2020.103721

Cited by 10 publications

(6 citation statements)

References 60 publications

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“…For the generation of the spectral library, DDA raw data were analyzed with Spectronaut software and a mass spectrometer vendor-independent software from Biognosys. 47 The DDA files were searched against the Swiss-Prot human database (20 353 entries released on Aug 10, 2020) and the decoy proteins were produced from pseudoreversed sequences of the target proteins. The digestion of trypsin was allowed with a maximum of two missed cleavages.…”

Section: ■ Materials and Methodsmentioning

confidence: 99%

Serum-Derived Exosomal Proteins as Potential Candidate Biomarkers for Hepatocellular Carcinoma

et al. 2021

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Hepatocellular carcinoma (HCC) is the most common form of hepatic malignancies. The diagnosis of HCC remains challenging due to the low sensitivity and specificity of the diagnostic method. Exosomes, which are abundant in various proteins from parent cells, play pivotal roles in intercellular communication and have been confirmed as promising sources of disease biomarkers. Herein, we performed a simple but robust proteomic profiling on exosomes derived from 1 μL of serum using a data-independent acquisition (DIA) method for the first time, to screen potential biomarkers for the diagnosis of HCC. Ten pivotal differentially expressed proteins (DEPs) (von Willebrand factor (VWF), LGALS3BP, TGFB1, SERPINC1, HPX, HP, HBA1, FGA, FGG, and FGB) were screened as a potential candidate biomarker panel, which could completely discriminate patients with HCC from normal control (NC). Interestingly, Gene Expression Profiling Interactive Analysis (GEPIA) revealed that the expression levels of four genes increased and those of six genes decreased in HCC tissues compared with normal tissues, which were in concordance with protein expression levels. In conclusion, we screened 10 exosomal proteins holding promise for acting as a potential candidate biomarker panel for detection of HCC through a simple but robust proteomic profiling.

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Section: ■ Materials and Methodsmentioning

confidence: 99%

Serum-Derived Exosomal Proteins as Potential Candidate Biomarkers for Hepatocellular Carcinoma

et al. 2021

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“…This leads to accurate peptide quantification with no limitation to profiling predefined interested peptides [121]. Rappsilber group applied DIA to QCLMS and found that DIA was indeed able to improve QCLMS[37, 122]. For a broader application of DIA in the future, development of robust data analysis tools for efficient processing of the highly convoluted spectral data is required.…”

Section: Progress In Experimental Methodologiesmentioning

confidence: 99%

Progress in methodologies and quality‐control strategies in protein cross‐linking mass spectrometry

et al. 2021

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Deciphering the interaction networks and structural dynamics of proteins is pivotal to better understanding their biological functions. Cross‐linking mass spectrometry (XL‐MS) is a powerful and increasingly popular technology that provides information about protein‐protein interactions and their structural constraints for individual proteins and multiprotein complexes on a proteome‐scale. In this review, we first assess the coverage and depth of the XL‐MS technique by utilizing publicly available datasets. We then delve into the progress in XL‐MS experimental and computational methodologies and examine different quality‐control strategies reported in the literature. Finally, we discuss the progress in XL‐MS applications along with the scope for future improvements.

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“…Nowadays, it is possible to take advantage of different strategies to obtain QXL-MS data, such as isotope-labeled cross-linkers, metabolic or isobaric labeling (Figure ), label-free data-dependent acquisition (DDA), or data-independent acquisition (DIA) modes. ,, …”

Section: Quantitative Cross-linkingmentioning

confidence: 99%

Cross-Linking Mass Spectrometry for Investigating Protein Conformations and Protein–Protein Interactions─A Method for All Seasons

Piersimoni¹,

Kastritis

Arlt³

et al. 2021

Chem. Rev.

168

103

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Mass spectrometry (MS) has become one of the key technologies of structural biology. In this review, the contributions of chemical cross-linking combined with mass spectrometry (XL-MS) for studying three-dimensional structures of proteins and for investigating protein–protein interactions are outlined. We summarize the most important cross-linking reagents, software tools, and XL-MS workflows and highlight prominent examples for characterizing proteins, their assemblies, and interaction networks in vitro and in vivo. Computational modeling plays a crucial role in deriving 3D-structural information from XL-MS data. Integrating XL-MS with other techniques of structural biology, such as cryo-electron microscopy, has been successful in addressing biological questions that to date could not be answered. XL-MS is therefore expected to play an increasingly important role in structural biology in the future.

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A protocol for studying structural dynamics of proteins by quantitative crosslinking mass spectrometry and data-independent acquisition

Cited by 10 publications

References 60 publications

Serum-Derived Exosomal Proteins as Potential Candidate Biomarkers for Hepatocellular Carcinoma

Serum-Derived Exosomal Proteins as Potential Candidate Biomarkers for Hepatocellular Carcinoma

Progress in methodologies and quality‐control strategies in protein cross‐linking mass spectrometry

Cross-Linking Mass Spectrometry for Investigating Protein Conformations and Protein–Protein Interactions─A Method for All Seasons

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