pFind Studio: a computational solution for mass spectrometry-based proteomics
2023
APPLIED SCIENCES-BASEL2023. Xiang Zhang et al.
Shandong Univ Technol, Sch Comp Sci & Technol, Zibo 255049, Peoples R China
ABSTRACT:Although data-independent acquisition (DIA) has the ability to identify and quantify all peptides in a sample, highly complex mixed mass spectra present difficulties for accurate peptide and protein identification. Additionally, the correspondence between the precursor and its fragments is broken, making it challenging to perform peptide identification directly using conventional DDA search engines. In this paper, we propose a cosine-similarity-based deconvolution method: CorrDIA. This is achieved by reconstructing the correspondence between precursor and fragment ions based on the consistency of extracted ion chromatograms (XICs). A deisotope peak cluster operation is added and centered on the MS/MS spectrum to improve the accuracy of spectrum interpretation and increase the number of identified peptides. The resulting MS/MS spectra can be identified using any data-dependent acquisition (DDA) sequencing software. The experimental results demonstrate that the number of peptide results increased by 12 percent and 21 percent respectively, and the repetition rate decreased by 12 percent. This reduces mass spectra complexity and difficulties in mass spectra analysis without the need for any mass spectra libraries.
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The Journal of biological chemistry2023. Yan, Sheng et al.
Beijing Key Laboratory of DNA Damage Response and College of Life Sciences, Capital Normal University, Beijing, China
ABSTRACT:The O-linked beta-N-acetylglucosamine (O-GlcNAc) transferase (OGT) mediates intracellular O-GlcNAcylation modification. O-GlcNAcylation occurs on Ser/Thr residues and is important for numerous physiological processes. OGT is essential for dividing mammalian cells and is involved in many human diseases; however, many of its fundamental substrates during cell division remain unknown. Here, we focus on the effect of OGT on polo-like kinase 1 (PLK1), a mitotic master kinase that governs DNA replication, mitotic entry, chromosome segregation, and mitotic exit. We show that PLK1 interacts with OGT and is O-GlcNAcylated. By utilizing stepped collisional energy/higher-energy collisional dissociation mass spectrometry, we found a peptide fragment of PLK1 that is modified by O-GlcNAc. Further mutation analysis of PLK1 shows that the T291A mutant decreases O-GlcNAcylation. Interestingly, T291N is a uterine carcinoma mutant in The Cancer Genome Atlas. Our biochemical assays demonstrate that T291A and T291N both increase PLK1 stability. Using stable H2B-GFP cells, we found that PLK1-T291A and PLK1-T291N mutants display chromosome segregation defects and result in misaligned and lagging chromosomes. In mouse xenograft models, we demonstrate that the O-GlcNAc-deficient PLK1-T291A and PLK1-T291N mutants enhance uterine carcinoma in animals. Hence, we propose that OGT partially exerts its mitotic function through O-GlcNAcylation of PLK1, which might be one mechanism by which elevated levels of O-GlcNAc promote tumorigenesis.
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Nature Communications2023. Hayes, Brooke K et al.
Biomedicine Discovery Institute, Department of Microbiology, Monash University, Clayton Melbourne, VIC,Australia
ABSTRACT:
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Antioxidants2023. Zhou, Xiangzhe et al.
School of Life Science, Beijing Institute of Technology, Beijing 100081, China
ABSTRACT:
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Journal of Agricultural and Food Chemistry2023. Ying Zhang et al.
Nanchang Univ, Sino German Joint Res Inst, Nanchang 330047, Peoples R China; Nanchang Univ, State Key Lab Food Sci & Technol, Nanchang 330047, Peoples R China
ABSTRACT:Giventhat roasting changes the structure and allergenicity ofpeanut allergens, the structural information of peanut allergens mustbe expounded to explain the alteration in their allergenicity. Thiswork focused on allergen aggregations (AAs) in roasted peanuts. IgErecognition capability was assessed via western blot analysis. Thedisulfide bond (DB) rearrangement and chemical modification in AAswere identified by combining mass spectroscopy and software tools,and structural changes induced by cross-links were displayed by moleculardynamics and PyMOL software. Results showed that AAs were stronglyrecognized by IgE and cross-linked mainly by DBs. The types of DBrearrangement in AAs included interprotein (98 peptide pairs), intraprotein(22 peptide pairs), and loop-linked (6 peptides) DBs. Among allergens,Ara h 2 and Ara h 6 presented the most cysteine residues to cross-linkfwith others or themselves. DB rearrangement involved IgE epitopesand induced structural changes. Ara h 1 and Ara h 3 were predominantlychemically modified. Moreover, chemical modification altered the localstructures of proteins, which may change the allergenic potentialof allergens.
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Nature Cancer2023. Kamal Manda et al.
Arun P. Wiita
ABSTRACT:Safely expanding indications for cellular therapies has been challenging given a lack of highly cancer-specific surface markers. Here we explore the hypothesis that tumor cells express cancer-specific surface protein conformations that are invisible to standard target discovery pipelines evaluating gene or protein expression, and these conformations can be identified and immunotherapeutically targeted. We term this strategy integrating cross-linking mass spectrometry with glycoprotein surface capture 'structural surfaceomics'. As a proof of principle, we apply this technology to acute myeloid leukemia (AML), a hematologic malignancy with dismal outcomes and no known optimal immunotherapy target. We identify the activated conformation of integrin beta(2) as a structurally defined, widely expressed AML-specific target. We develop and characterize recombinant antibodies to this protein conformation and show that chimeric antigen receptor T cells eliminate AML cells and patient-derived xenografts without notable toxicity toward normal hematopoietic cells. Our findings validate an AML conformation-specific target antigen and demonstrate a tool kit for applying these strategies more broadly.
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Nature Communications2023. Gautham Yepuri et al.
Diabetes Research Program, Division of Endocrinology, Diabetes and Metabolism, Department of Medicine, NYU Grossman School of Medicine, New York, New York, 10016, USA
ABSTRACT:Inter-organelle contact and communication between mitochondria and sarco/endoplasmic reticulum (SR/ER) maintain cellular homeostasis and are profoundly disturbed during tissue ischemia. We tested the hypothesis that the formin Diaphanous-1 (DIAPH1), which regulates actin dynamics, signal transduction and metabolic functions, contributes to these processes. We demonstrate that DIAPH1 interacts directly with Mitofusin-2 (MFN2) to shorten mitochondria-SR/ER distance, thereby enhancing mitochondria-ER contact in cells including cardiomyocytes, endothelial cells and macrophages. Solution structure studies affirm the interaction between the Diaphanous Inhibitory Domain and the cytosolic GTPase domain of MFN2. In male rodent and human cardiomyocytes, DIAPH1-MFN2 interaction regulates mitochondrial turnover, mitophagy, and oxidative stress. Introduction of synthetic linker construct, which shorten the mitochondria-SR/ER distance, mitigated the molecular and functional benefits of DIAPH1 silencing in ischemia. This work establishes fundamental roles for DIAPH1-MFN2 interaction in the regulation of mitochondria-SR/ER contact networks. We propose that targeting pathways that regulate DIAPH1-MFN2 interactions may facilitate recovery from tissue ischemia.
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Science Advances2023. Li Jie et al.
Department of Biochemistry and Molecular Pharmacology, NewYork University Grossman School of Medicine, NewYork, NY 10016, USA.
ABSTRACT:Intracellular degradation of proteins and organelles by the autophagy-lysosome system is essential for cellular quality control and energy homeostasis. Besides degradation, endolysosomal organelles can fuse with the plasma membrane and contribute to unconventional secretion. Here, we identify a function for mammalian SKP1 in endolysosomes that is independent of its established role as an essential component of the family of SCF/CRL1 ubiquitin ligases. We found that, under nutrient-poor conditions, SKP1 is phosphorylated on Thr(131), allowing its interaction with V-1 subunits of the vacuolar ATPase (V-ATPase). This event, in turn, promotes V-ATPase assembly to acidify late endosomes and enhance endolysosomal degradation. Under nutrient-rich conditions, SUMOylation of phosphorylated SKP1 allows its binding to and dephosphorylation by the PPM1B phosphatase. Dephosphorylated SKP1 interacts with SEC22B to promote unconventional secretion of the content of less acidified hybrid endosomal/autophagic compartments. Collectively, our study implicates SKP1 phosphorylation as a switch between autophagy and unconventional secretion in a manner dependent on cellular nutrient status.
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International Journal of Molecular Sciences2023. Hana Popelka et al.
Life Sciences Institute, University of Michigan, Ann Arbor, MI 48109, USA
Department of Molecular, Cellular and Developmental Biology, University of Michigan, Ann Arbor, MI 48109, USA
Department of Pathology, Michigan Medicine, University of Michigan, Ann Arbor, MI 48109, USA
ABSTRACT:The Atg12 protein in yeast is an indispensable polypeptide in the highly conserved ubiquitin-like conjugation system operating in the macroautophagy/autophagy pathway. Atg12 is covalently conjugated to Atg5 through the action of Atg7 and Atg10; the Atg12-Atg5 conjugate binds Atg16 to form an E3 ligase that functions in a separate conjugation pathway involving Atg8. Atg12 is comprised of a ubiquitin-like (UBL) domain preceded at the N terminus by an intrinsically disordered protein region (IDPR), a domain that comprises a major portion of the protein but remains elusive in its conformation and function. Here, we show that the IDPR in unconjugated Atg12 is positioned in proximity to the UBL domain, a configuration that is important for the functional structure of the protein. A major deletion in the IDPR disrupts intactness of the UBL domain at the unconjugated C terminus, and a mutation in the predicted alpha 0 helix in the IDPR prevents Atg12 from binding to Atg7 and Atg10, which ultimately affects the protein function in the ubiquitin-like conjugation cascade. These findings provide evidence that the IDPR is an indispensable part of the Atg12 protein from yeast.
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FEBS Open bio2023. Huang Peng et al.
Department of Experimental Medical Science, Lund University, Sweden
ABSTRACT:Glucose transporters (GLUTs) are responsible for transporting hexose molecules across cellular membranes. In adipocytes, insulin stimulates glucose uptake by redistributing GLUT4 to the plasma membrane. In unstimulated adipose-like mouse cell lines, GLUT4 is known to be retained intracellularly by binding to TUG protein, while upon insulin stimulation, GLUT4 dissociates from TUG. Here, we report that the TUG homolog in human, ASPL, exerts similar properties, i.e., forms a complex with GLUT4. We describe the structural details of complex formation by combining biochemical assays with cross-linking mass spectrometry and computational modeling. Combined, the data suggest that the intracellular domain of GLUT4 binds to the helical lariat of ASPL and contributes to the regulation of GLUT4 trafficking by cooperative binding.
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