Article
Pharmacology & Pharmacy
Ying Zhang, Qinghua Cai, Yuxiang Luo, Yu Zhang, Huilin Li
Summary: Defects in ribosomal proteins are linked to various diseases, and comprehensive characterization of ribosomal proteoforms is important for discovering potential disease biomarkers or targets. The combination of top-down and bottom-up proteomics approaches successfully identified and analyzed the different forms and modifications of ribosomal proteins.
JOURNAL OF PHARMACEUTICAL ANALYSIS
(2023)
Article
Multidisciplinary Sciences
Jinwei Xing, Elizabeth R. Chrastil, Douglas A. Nitz, Jeffrey L. Krichmar
Summary: Humans and animals have the ability to translate their position from one spatial frame of reference to another, and seamlessly switch between top-down and first-person views. The medial temporal lobe and other cortical regions are found to contribute to this function. By using variational autoencoders to reconstruct views, researchers gain insights into how the neural system carries out these computations.
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA
(2022)
Article
Biochemical Research Methods
Georg J. Pirklbauer, Christian E. Stieger, Manuel Matzinger, Stephan Winkler, Karl Mechtler, Viktoria Dorfer
Summary: Cross-linking mass spectrometry (XL-MS) has enabled insights into protein structures and interactions, with cleavable cross-linkers further reducing search space and promoting proteome-wide studies. However, some search engines struggle with validating identified cross-linked peptides and estimating false discovery rates (FDR). The new search engine MS Annika provides realistic FDR estimates without the need for arbitrary score cutoffs, offering on average 44% more identifications at a similar or better true FDR compared to other tools.
JOURNAL OF PROTEOME RESEARCH
(2021)
Article
Chemistry, Analytical
Kyle A. Brown, Morgan K. Gugger, Zhen Yu, David Moreno, Song Jin, Ying Ge
Summary: Nonionic surfactants are commonly used reagents for cell lysis and protein extraction in structural biology. However, their presence often interferes with protein analysis by electrospray ionization-mass spectrometry (ESI-MS). This study introduces a cleavable surfactant, DSSM, which is compatible with ESI-MS analysis and enables top-down proteomics characterization. DSSM can replace DDM in proteomic experiments and structural biology studies.
ANALYTICAL CHEMISTRY
(2023)
Article
Chemistry, Analytical
Kyle A. Brown, Morgan K. Gugger, Zhen Yu, David Moreno, Song Jin, Ying Ge
Summary: Nonionic surfactants are widely used in cell lysis for protein extraction, stabilization, and purification in structural biology. However, they often interfere with electrospray ionization-mass spectrometry (ESI-MS) analysis. In this study, a cleavable nonionic surfactant, n-decyl-disulfide-beta-D-maltoside (DSSM), was developed to overcome this limitation. DSSM is compatible with ESI-MS and reversed-phase liquid chromatography-MS analysis and allows for characterization of membrane proteins and endogenous proteins, making it a potential replacement for the commonly used surfactant DDM in proteomic experiments and structural biology studies.
ANALYTICAL CHEMISTRY
(2023)
Article
Chemistry, Analytical
Francis Berthias, Hayden A. Thurman, Gayani Wijegunawardena, Haifan Wu, Alexandre A. Shvartsburg, Ole N. Jensen
Summary: Advances in proteomics have shown the ubiquity and biological importance of diverse proteoforms. The ability to separate and analyze intact isomeric proteoforms is crucial for understanding their functions. A novel technique using trapped IMS was demonstrated to achieve baseline resolution of isomeric proteoforms, opening new avenues in proteomics and epigenetics.
ANALYTICAL CHEMISTRY
(2022)
Article
Biochemical Research Methods
Yong Cao, Xin-Tong Liu, Peng-Zhi Mao, Zhen-Lin Chen, Ching Tarn, Meng-Qiu Dong
Summary: In mass spectrometry data analysis, different search engines have different settings for cross-linkable sites. Some only include lysine and protein N-terminus, while others also include serine, threonine, and tyrosine by default. By using chemically inert amino acids as cross-linkable sites, we show that STY-cross-links identified by software are as reliable as GVL-cross-links.
JOURNAL OF PROTEOME RESEARCH
(2023)
Article
Biochemical Research Methods
Lei Lu, Mark Scalf, Michael R. Shortreed, Lloyd M. Smith
Summary: Mesh is a novel dissociation strategy that can dissociate multiple charge states of a single proteoform with different collision energies, improving fragment ion sequence coverage and identification ratios in top-down proteomic analyses of complex samples. The strategy is implemented within the open-source instrument control software program MetaDrive, which performs real-time deconvolution and precursor selection.
JOURNAL OF THE AMERICAN SOCIETY FOR MASS SPECTROMETRY
(2021)
Article
Biochemical Research Methods
Andrea M. Brunner, Philip Lossl, Paul P. Geurink, Huib Ovaa, P. Albanese, A. F. Maarten Altelaar, Albert J. R. Heck, Richard A. Scheltema
Summary: Top-down proteomics combined with bioinformatics approaches can improve the localization and quantification of protein modifications, even for positional isomers with the same number of modifications at different sites. This method shows promising results in characterizing multiple modified proteins and estimating the stoichiometry of phosphorylated sites.
MOLECULAR & CELLULAR PROTEOMICS
(2021)
Article
Biochemical Research Methods
Konrad Winkels, Tomas Koudelka, Philipp T. Kaulich, Matthias Leippe, Andreas Tholey
Summary: Top-down proteomics (TDP) can provide additional information about post-translational modifications and C-termini. In this study, TDP workflows were used to analyze the proteome of Caenorhabditis elegans, and the identified proteoforms were validated using a bottom-up proteomics (BUP) approach. The results showed low overlap between the two methods, with caveats in both workflows contributing to this. The study also demonstrated the applicability of reductive dimethylation in TDP to confirm biological N-termini.
JOURNAL OF PROTEOME RESEARCH
(2022)
Article
Chemistry, Analytical
Charlotte Mase, Julien F. Maillard, Marco Piparo, Lukas Friederici, Christopher P. Rueger, Sabrina Marceau, Benoit Paupy, Marie Hubert-Roux, Carlos Afonso, Pierre Giusti
Summary: Pyrolysis is a promising method for converting plastic waste into valuable resources, but undesirable species generated during this process pose challenges for downstream upgrading. Advanced analytical instrumentation is needed to accurately and completely characterize the high molecular diversity found in these samples. The combination of gas chromatography with Fourier transform ion cyclotron resonance mass spectrometry, using different dopants for selective ionization, allows for better characterization of the complex organic mixtures in plastic pyrolysis oil.
Article
Chemistry, Applied
Mengxue Fang, Xiaoying Luo, Shanbai Xiong, Tao Yin, Yang Hu, Ru Liu, Hongying Du, Youming Liu, Juan You
Summary: Increasing the cross-linking degree of surimi gels results in a higher number of cross-linking sites, a shift in the main possible cross-linking domain, and changes in digestion sites. The study highlights the complex quantity and location relationships between cross-linking and digestion.
Article
Engineering, Environmental
Estefania Alvarez-Castillo, Maria Jesus Pelagio, Carlos Bengoechea, Antonio Guerrero
Summary: This study aims to improve the mechanical properties of superabsorbent bioplastics based on porcine plasma protein by adding different amounts of glutaraldehyde as a cross-linking agent. The results showed that adding glutaraldehyde through immersion can significantly reduce soluble material loss.
JOURNAL OF ENVIRONMENTAL CHEMICAL ENGINEERING
(2021)
Article
Chemistry, Multidisciplinary
Liju Xu, Yan Qiao, Dong Qiu
Summary: An adaptable crystal-domain cross-linking design is used to overcome the trade-off between stiffness and toughness in hydrogels, allowing for the simultaneous improvement of both properties. This finding provides a practical and universal method for designing stiff and tough hydrogels and has a profound impact on various applications requiring hydrogels with combined mechanical properties.
ADVANCED MATERIALS
(2023)
Article
Biochemical Research Methods
Emily A. Chapman, Timothy J. Aballo, Jake A. Melby, Tianhua Zhou, Scott J. Price, Kalina J. Rossler, Ienglam Lei, Paul C. Tang, Ying Ge
Summary: This study investigated the changes in sarcomeric proteins in the myocardia of ischemic cardiomyopathy (ICM) patients using liquid chromatography-mass spectrometry (LC-MS). The results revealed altered phosphorylation and expression of several sarcomeric proteins in ICM hearts, indicating severe dysregulation of the contractile apparatus.
JOURNAL OF PROTEOME RESEARCH
(2023)
Article
Biochemistry & Molecular Biology
Ruzena Filandrova, Karel Valis, Jiri Cermy, Josef Chmelik, Lukas Slavata, Jan Fiala, Michal Rosulek, Daniel Kavan, Petr Man, Tomas Chum, Marek Cebecauer, Daniele Fabris, Petr Novak
Summary: TEAD transcription factors regulate gene expression through interactions with DNA and other proteins. Research findings suggest that TEAD1 can bind to the inverted version of the M-CAT motif, and that inverted motifs in the human genome may have biological relevance.
Article
Allergy
Veronika Kanderova, Tamara Svobodova, Simon Borna, Martina Fejtkova, Vendula Martinu, Jana Paderova, Michael Svaton, Jarmila Kralova, Eva Fronkova, Adam Klocperk, Stepanka Pruhova, Min Ae Lee-Kirsch, Ludmila Hornofova, Miroslav Koblizek, Petr Novak, Olga Zimmermannova, Zuzana Parackova, Anna Sediva, Tomas Kalina, Ales Janda, Jana Kayserova, Marcela Dvorakova, Milan Macek, Petr Pohunek, Petr Sedlacek, Ashleigh Poh, Matthias Ernst, Tomas Brdicka, Ondrej Hrusak, Jan Lebl
Summary: This article reports a new monogenic autoinflammatory disorder caused by a de novo activating mutation in the HCK gene, characterized by cutaneous vasculitis and chronic pulmonary inflammation. The study shows the aberrant functions of mutant HCK and the therapeutic efficacy of the Janus kinase 1/2 inhibitor Ruxolitinib in treating inflammatory lung disease.
JOURNAL OF ALLERGY AND CLINICAL IMMUNOLOGY
(2022)
Article
Chemistry, Analytical
Marek Polak, Ghazaleh Yassaghi, Daniel Kavan, Frantisek Filandr, Jan Fiala, Zdenek Kukacka, Petr Halada, Dmitry S. Loginov, Petr Novak
Summary: The combination of covalent labeling techniques with mass spectrometry is a progressive approach for studying protein surfaces and protein-ligand interactions. In this study, the interaction interface between the DNA binding domain of the FOXO4 protein and the DNA binding element was mapped. Different approaches were used to confirm the results and compare with other studies, showing consistent findings. This method is important for understanding the structure and dynamics of protein-DNA interactions.
ANALYTICAL CHEMISTRY
(2022)
Article
Biochemistry & Molecular Biology
Dmitry S. Loginov, Jan Fiala, Peter Brechlin, Gary Kruppa, Petr Novak
Summary: Structural mass spectrometry methods, particularly FPOP, are widely used for studying protein structure and dynamics. In this study, FPOP analysis of the Hb-Hp complex revealed specific residues affected by the complex formation and provided insights into the interacting amino acids. The data, available via ProteomeXchange, can be further utilized for investigating protein-protein interactions.
BIOCHIMICA ET BIOPHYSICA ACTA-PROTEINS AND PROTEOMICS
(2022)
Article
Biochemistry & Molecular Biology
Valerio G. Giacobelli, Kosuke Fujishima, Martin Lepsik, Vyacheslav Tretyachenko, Tereza Kadava, Mikhail Makarov, Lucie Bednarova, Petr Novak, Klara Hlouchova
Summary: RNA-peptide/protein interactions are crucial to life and have been studied in the context of early evolution. This study focused on an RNA-binding variant of the ribosomal uL11 C-terminal domain, selected from a library of prebiotically plausible amino acid sequences. The study found that the selected variant binds RNA with a similar overall affinity, utilizing ion bridging interactions for stabilization in the absence of aromatic/basic residues. These findings provide insights into how early protein-RNA interactions may have evolved.
MOLECULAR BIOLOGY AND EVOLUTION
(2022)
Article
Chemistry, Analytical
Ghazaleh Yassaghi, Zdenek Kukacka, Jan Fiala, Daniel Kavan, Petr Halada, Michael Volny, Petr Novak
Summary: Fast photochemical oxidation of proteins (FPOP) footprinting is a method that uses chemical reactions to label protein structures. It provides information about protein conformation, dynamics, and interactions. FPOP samples can be analyzed using either bottom-up or top-down mass spectrometry approaches. The top-down approach allows for more specific selection of protein subpopulations, but is limited to smaller proteins. Comparison with bottom-up data confirms the usefulness of the top-down approach in oxidative protein footprinting.
ANALYTICAL CHEMISTRY
(2022)
Meeting Abstract
Hematology
Natividad Alquezar-Artieda, Jennie Roberts, Daniela Kuzilkova, Katerina Hlozkova, Alena Pecinova, Petr Pecina, Martina Zwyrtkova, Eliska Potuckova, Daniel Kavan, Ivana Hermanova, Marketa Zaliova, Petr Novak, Tomas Mracek, Daniel A. Tennant, Jan Trka, Julia Starkova
Article
Multidisciplinary Sciences
Vendula Studentova, Vendula Sudova, Ibrahim Bitar, Veronika Paskova, Jiri Moravec, Petr Pompach, Michael Volny, Petr Novak, Jaroslav Hrabak
Summary: This study focused on a deep analysis of OXA-48-like-producing Enterobacterales, especially strains showing poor hydrolytic activity. A new alternative way of antibiotic modification by OXA-48-like enzymes was identified. Various methods were used to determine the carbapenemase activity, and it was found that meropenem-derived beta-lactone was preferentially produced by strains that showed false-negative results using Carba NP test.
SCIENTIFIC REPORTS
(2022)
Letter
Hematology
Natividad Alquezar-Artieda, Daniela Kuzilkova, Jennie Roberts, Katerina Hlozkova, Alena Pecinova, Petr Pecina, Martina Zwyrtkova, Eliska Potuckova, Daniel Kavan, Ivana Hermanova, Marketa Zaliova, Petr Novak, Tomas Mracek, Lucie Sramkova, Daniel A. Tennant, Jan Trka, Julia Starkova
Article
Multidisciplinary Sciences
Pieter-Jan Vermeire, Anastasia V. Lilina, Hani M. Hashim, Lada Dlabolova, Jan Fiala, Steven Beelen, Zdenek Kukacka, Jeremy N. Harvey, Petr Novak, Sergei V. Strelkov
Summary: Intermediate filaments (IFs) are essential components of the metazoan cytoskeleton and can self-assemble from soluble tetrameric species into typical 10-12 nm wide filaments. The molecular structure of mature IFs is poorly understood, but this study reveals that the formation of tetramers is dependent on the presence of the coil1A domain and is synergistically stabilized by interactions between coil1B domains, coil1A, coil2, and the head and rod domains. Additionally, small-angle X-ray scattering experiments show that elongated A(11) tetramers are dominant in low ionic strength solutions with structural flexibility in the terminal domains.
SCIENTIFIC REPORTS
(2023)
Article
Biochemical Research Methods
Josef Dvorak, Jana Novakova, Lucie Kraftova, Vendula Studentova, Martin Matejovic, Jaroslav Radej, Thomas Karvunidis, Jan Horak, Marcela Kralovcova, Jaroslav Hrabak, Zuzana Kalaninova, Michael Volny, Petr Novak, Petr Pompach
Summary: A rapid PCT detection method using MALDI-TOF mass spectrometry was developed and validated against LC-MS bottom-up workflow. The new method detects intact PCT and uncovers its alterations in septic patients' serum.
CLINICAL PROTEOMICS
(2023)
Article
Ecology
Brennen Heames, Filip Buchel, Margaux Aubel, Vyacheslav Tretyachenko, Dmitry Loginov, Petr Novak, Andreas Lange, Erich Bornberg-Bauer, Klara Hlouchova
Summary: A comparison between de novo genes and synthetic random sequences reveals no difference in biophysical properties, but in vitro expression demonstrates de novo proteins' higher solubility, indicating their better integration into the cellular system. The emergence of de novo genes provides a pathway for the formation of new proteins from previously non-coding DNA. While the likelihood of assuming a soluble and stable tertiary structure in de novo proteins remains unclear, evidence suggests that native-like biophysical properties are abundant in sequence space. Experimental characterization of putative de novo proteins in human and fly shows that their solubility and structure propensity are moderately higher compared to synthetic random proteins. The DnaK chaperone system further enhances the solubility of de novo proteins. Therefore, de novo proteins may be better integrated into the cellular system than random sequences due to their higher solubility.
NATURE ECOLOGY & EVOLUTION
(2023)
Article
Chemistry, Analytical
Marek Polak, Michael Palasser, Alan Kadek, Daniel Kavan, Christopher A. Wootton, Marc-Andre Delsuc, Kathrin Breuker, Petr Novak, Maria A. van Agthoven
Summary: Two-dimensional mass spectrometry (2D MS) is a multiplexed tandem mass spectrometry method that uses modulation of precursor ion radii to show the fragmentation patterns of analytes. This study introduces quadrupolar detection in 2D MS and discusses its advantages, as well as adapts existing data processing techniques for accurate frequency-to-mass conversion. The method is applied to the top-down analysis of covalently labeled ubiquitin and the label-free relative quantification of biomolecule isoforms.
ANALYTICAL CHEMISTRY
(2023)
Article
Biochemical Research Methods
Thomas Lambert, Marius Gramlich, Luisa Stutzke, Luke Smith, Dingyu Deng, Philipp D. Kaiser, Ulrich Rothbauer, Justin L. P. Benesch, Cornelia Wagner, Maximiliane Koenig, Petr Pompach, Petr Novak, Anne Zeck, Kasper D. Rand
Summary: Protein glycosylation is a common protein modification, but determining the HDX of glycosylated protein segments has been challenging. In this study, we developed a column containing PNGase Rc that can be integrated into a conventional HDX-MS setup to improve glycoprotein analysis. The PNGase Rc column enables efficient online removal of N-linked glycans and determination of glycosylated regions' HDX in complex glycoproteins. We also demonstrated the column's use in mapping the binding epitope of a mAb to c-Met, a complex glycoprotein drug target. Overall, our work shows that HDX-MS with the integrated PNGase Rc column allows fast and efficient deglycosylation under harsh quench conditions and provides comprehensive analysis of complex glycoproteins.
JOURNAL OF THE AMERICAN SOCIETY FOR MASS SPECTROMETRY
(2023)
Article
Biochemistry & Molecular Biology
Eliska Koutna, Vanda Lux, Tomas Kouba, Jana Skerlova, Jiri Novacek, Pavel Srb, Rozalie Hexnerova, Hana Svachova, Zdenek Kukacka, Petr Novak, Milan Fabry, Simon Poepsel, Vaclav Veverka
Summary: This study reports the ability of the PWWP domain of transcriptional coactivator LEDGF to bind to specific histone modifications and reveals the role of the intrinsically disordered region in nucleosome binding. The interplay between the H3K36 methylation reader and protein binding module of LEDGF facilitates productive transcription elongation.
NUCLEIC ACIDS RESEARCH
(2023)