scholarly journals Large-scale identification of protein histidine methylation in human cells

2021 ◽  
Author(s):  
Sebastian Kapell ◽  
Magnus E. Jakobsson
Keyword(s):  
Large Scale ◽  
Human Cells ◽  
System Level ◽  
Mass Spectrometry Analysis ◽  
Regulatory Function ◽  
Functional Importance ◽  
Spectrometry Analysis ◽  
Arginine Residues ◽  
Sequence Requirements ◽  
Level Analysis

ABSTRACTMethylation can occur on histidine, lysine and arginine residues in proteins and often serves a regulatory function. Histidine methylation has recently attracted notable attention through the discovery of the human histidine methyltransferase enzymes SETD3 and METTL9. There are currently no methods to enrich histidine methylated peptides for mass spectrometry analysis and large-scale analyses of the modification are hitherto absent. In the present study we query ultra-comprehensive proteomic datasets to generate a resource of histidine methylation sites in human cells. We use this resource to explore the frequency, localization, targeted domains, protein types and sequence requirements of histidine methylation and benchmark all analyses to methylation events on lysine and arginine. Our results demonstrate that histidine methylation is widespread in human cells and tissues and that the modification is over-represented in regions of mono-spaced histidine repeats. We also report colocalization of the modification with functionally important phosphorylation sites and disease associated mutations to identify regions of likely regulatory and functional importance. Taken together, we here report a system level analysis of human histidine methylation and our results represent a comprehensive resource enabling targeted studies of individual histidine methylation events.

scholarly journals Large-scale identification of protein histidine methylation in human cells

2021 ◽  
Vol 3 (2) ◽  
Author(s):  
Sebastian Kapell ◽  
Magnus E Jakobsson
Keyword(s):  
Large Scale ◽  
Human Cells ◽  
System Level ◽  
Mass Spectrometry Analysis ◽  
Regulatory Function ◽  
Lysine Methylation ◽  
Spectrometry Analysis ◽  
Arginine Residues ◽  
Sequence Requirements ◽  
Level Analysis

Abstract Methylation can occur on histidine, lysine and arginine residues in proteins and often serves a regulatory function. Histidine methylation has recently attracted attention through the discovery of the human histidine methyltransferase enzymes SETD3 and METTL9. There are currently no methods to enrich histidine methylated peptides for mass spectrometry analysis and large-scale studies of the modification are hitherto absent. Here, we query ultra-comprehensive human proteome datasets to generate a resource of histidine methylation sites. In HeLa cells alone, we report 299 histidine methylation sites as well as 895 lysine methylation events. We use this resource to explore the frequency, localization, targeted domains, protein types and sequence requirements of histidine methylation and benchmark all analyses to methylation events on lysine and arginine. Our results demonstrate that histidine methylation is widespread in human cells and tissues and that the modification is over-represented in regions of mono-spaced histidine repeats. We also report colocalization of the modification with functionally important phosphorylation sites and disease associated mutations to identify regions of likely regulatory and functional importance. Taken together, we here report a system level analysis of human histidine methylation and our results represent a comprehensive resource enabling targeted studies of individual histidine methylation events.

scholarly journals Co-Localization of Crotamine with Internal Membranes and Accentuated Accumulation in Tumor Cells

Molecules ◽  
2018 ◽  
Vol 23 (4) ◽  
pp. 968 ◽  
Author(s):  
Nicole Mambelli-Lisboa ◽  
Juliana Mozer Sciani ◽  
Alvaro Rossan Brandão Prieto da Silva ◽  
Irina Kerkis
Keyword(s):  
Molecular Mass ◽  
Tumor Cells ◽  
Large Scale ◽  
Time Lapse ◽  
Mass Spectrometry Analysis ◽  
Molecular Probe ◽  
Spastic Paralysis ◽  
Spectrometry Analysis ◽  
Hind Limbs ◽  
Wide Range

Crotamine is a highly cationic; cysteine rich, cross-linked, low molecular mass cell penetrating peptide (CPP) from the venom of the South American rattlesnake. Potential application of crotamine in biomedicine may require its large-scale purification. To overcome difficulties related with the purification of natural crotamine (nCrot) we aimed in the present study to synthesize and characterize a crotamine analog (sCrot) as well investigate its CPP activity. Mass spectrometry analysis demonstrates that sCrot and nCrot have equal molecular mass and biological function—the capacity to induce spastic paralysis in the hind limbs in mice. sCrot CPP activity was evaluated in a wide range of tumor and non-tumor cell tests performed at different time points. We demonstrate that sCrot-Cy3 showed distinct co-localization patterns with intracellular membranes inside the tumor and non-tumor cells. Time-lapse microscopy and quantification of sCrot-Cy3 fluorescence signalss in living tumor versus non-tumor cells revealed a significant statistical difference in the fluorescence intensity observed in tumor cells. These data suggest a possible use of sCrot as a molecular probe for tumor cells, as well as, for the selective delivery of anticancer molecules into these tumors.

Arginine Methylation of Runx1 Regulates Its Biological and Transcriptional Activities.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 2041-2041
Author(s):  
Xinyang Zhao ◽  
Animesh Parkanani ◽  
Jin Zhang ◽  
Richard Dunne ◽  
Andrew Xiao ◽  
...  
Keyword(s):  
Transcriptional Repression ◽  
Arginine Methylation ◽  
Serine Phosphorylation ◽  
Context Dependency ◽  
Mass Spectrometry Analysis ◽  
Runt Domain ◽  
Spectrometry Analysis ◽  
Gene Reporter Assay ◽  
Arginine Residues

Abstract The AML1/Runx1 protein is required for definitive hematopoiesis and for the maturation of adult megakaryocytic cells. Alterations of the Runx1 gene by mutations, deletions and chromosome translocations are associated with several types of acute leukemia. Runx1 functions as both an activator and repressor of gene transcription with promoter and cell type context dependency. Likely, this relates to the ability of Runx1 to interact with a variety of transcription factors such as MEF, C/EBPa, Ets-1 and GATA-1 and also with repressor proteins such as Groucho, mSin3 and HDACs. Runx1 is post-translationally modified through acetylation and phosphorylation, and the acetylable and phosphorylable forms of Runx1 can activate transcription to higher level in Runx1 dependent reporter assays when HATs or Erk2 are coexpressed. Runx1 has also been shown to be methylated on lysine residues by SUV39H1 methyltransferase in fibroblasts. Based on the presence of a SGRGK motif in the runt domain of Runx1, we have been examining whether Runx1 is methylated on arginine residues by the protein arginine methyltransferases (PRMT). We have found that PRMT1 and PRMT5 are associated with Runx1 in AML cells by co-immunoprecipitation assays and using in vitro by GST-pulldown assays with in vitro translated PRMT(s) have shown that the interactions are direct. Using a luciferase gene reporter assay, we show that PRMT1 acts synergistically with p300 to activate Runx1 mediated transcription in response to cell proliferation signals. We have mapped the arginine methylation sites in Runx1 using GST-Runx1 fusion proteins, site-specific mutagenesis and mass spectrometry analysis. We have found three potential arginine methylation sites, one in the Runt domain, and two in the Runx1 carboxy-terminal region. Interestingly, one of these sites is in the region shown to interact with both the mSIN3A transcriptional repression complex and with p300. This suggests that arginine methylation of Runx1 may affects its transcriptional activating and repressing functions. Chromatin immunoprecipitation assays are underway to show how arginine methylation of Runx1 affects its activities in hematopoietic cells. Additional studies examining the effects of cross-talk between arginine methylation, lysine acetylation and serine phosphorylation has on Runx1 functions (biological and biochemical) will be presented.

Application of IOT (Internet of Things) Technology in Chemical Testing with Large-Scale Analysis Instruments

2014 ◽  
Vol 701-702 ◽  
pp. 475-479
Author(s):  
Hong Bo Chen ◽  
Yi Zheng ◽  
Yan Gang Han
Keyword(s):  
Large Scale ◽  
Mass Spectrometry Analysis ◽  
Testing System ◽  
Spectrometry Analysis ◽  
Whole Process ◽  
Large Scale Analysis ◽  
Intelligent Methods ◽  
Internet Of Things Technology ◽  
Chemical Testing ◽  
Standard Development

In the field of chemical testing, it is of great importance to improve the accuracy and efficiency and reduce the risk caused by artificial factor with modern intelligent methods, which are critical to the standard development of testing labs. In this paper, intelligent control during the whole process of chromatography and mass spectrometry analysis with cloud computing technology was realized and discussed in detail. The intelligent testing system could be applied to the chemical analysis labs and spread to other fields.

Prediction and collection of protein–metabolite interactions

2021 ◽  
Author(s):  
Tianyi Zhao ◽  
Jinxin Liu ◽  
Xi Zeng ◽  
Wei Wang ◽  
Sheng Li ◽  
...  
Keyword(s):  
Small Molecule ◽  
Protein Interactions ◽  
Large Scale ◽  
Rapid Development ◽  
Area Under The Curve ◽  
Membrane Receptors ◽  
Mass Spectrometry Analysis ◽  
Spectrometry Analysis ◽  
Cellular Processes ◽  
The Rich

Abstract Interactions between proteins and small molecule metabolites play vital roles in regulating protein functions and controlling various cellular processes. The activities of metabolic enzymes, transcription factors, transporters and membrane receptors can all be mediated through protein–metabolite interactions (PMIs). Compared with the rich knowledge of protein–protein interactions, little is known about PMIs. To the best of our knowledge, no existing database has been developed for collecting PMIs. The recent rapid development of large-scale mass spectrometry analysis of biomolecules has led to the discovery of large amounts of PMIs. Therefore, we developed the PMI-DB to provide a comprehensive and accurate resource of PMIs. A total of 49 785 entries were manually collected in the PMI-DB, corresponding to 23 small molecule metabolites, 9631 proteins and 4 species. Unlike other databases that only provide positive samples, the PMI-DB provides non-interaction between proteins and metabolites, which not only reduces the experimental cost for biological experimenters but also facilitates the construction of more accurate algorithms for researchers using machine learning. To show the convenience of the PMI-DB, we developed a deep learning-based method to predict PMIs in the PMI-DB and compared it with several methods. The experimental results show that the area under the curve and area under the precision-recall curve of our method are 0.88 and 0.95, respectively. Overall, the PMI-DB provides a user-friendly interface for browsing the biological functions of metabolites/proteins of interest, and experimental techniques for identifying PMIs in different species, which provides important support for furthering the understanding of cellular processes. The PMI-DB is freely accessible at http://easybioai.com/PMIDB.

Proteomic analysis reveals the negative modulator of sperm function glycodelin as over-represented in semen exosomes isolated from asthenozoospermic patients

2019 ◽  
Vol 34 (8) ◽  
pp. 1416-1427 ◽  
Author(s):  
Valentina Murdica ◽  
Greta Chiara Cermisoni ◽  
Natasa Zarovni ◽  
Andrea Salonia ◽  
Paola Viganò ◽  
...  
Keyword(s):  
Western Blot ◽  
Proteomic Analysis ◽  
Large Scale ◽  
Reproductive Tract ◽  
Secretory Protein ◽  
Differentially Expressed ◽  
Mass Spectrometry Analysis ◽  
Inhibitory Protein ◽  
Reproductive Process ◽  
Spectrometry Analysis

ABSTRACT STUDY QUESTION Are there differences in the proteomic profile of exosomes isolated from seminal plasma of normozoospermic (NSP) and severe asthenozoospermic (SA) men, potentially contributing to sperm features? SUMMARY ANSWER A relevant group of proteins known to positively regulate sperm functions were over-represented in seminal exosomes of NSP men, i.e. cysteine-rich secretory protein-1 (CRISP1), while the inhibitory protein glycodelin was enriched in exosomes of SA subjects. WHAT IS KNOWN ALREADY Exosomes are secreted along the male reproductive tract and are thought to be involved in spermatozoa maturation and function. Ejaculated spermatozoa are still able to capture exosomes; exosomes of NSP individuals improve sperm motility and prompt capacitation, while exosomes of SA men fail to exert similar features. STUDY DESIGN, SIZE, DURATION Semen samples from NSP and SA men, aged 18 to 55 and registered at a single IVF center, were considered for this study project. Subjects were subdivided into three groups: a discovery cohort (five NSP men and six SA patients), a validation cohort (seven NSP and seven SA men) and the ‘glycodelin analysis’ cohort (20 NSP and 37 SA men). Exosomes were purified from semen of every participant. PARTICIPANTS/MATERIALS, SETTING, METHODS Exosomes were characterized by nanoparticle tracking analysis, transmission electron microscopy and western blot. Comprehensive proteomics analysis of the exosomal proteome was performed by nanoscale liquid chromatographic tandem mass spectrometry analysis. Funrich software was used to determine statistical enrichment of pathways, networks and Gene Ontology terms of the identified proteins. Validation of differentially expressed proteins was performed through ELISA and western blot analysis. MAIN RESULTS AND THE ROLE OF CHANCE The comprehensive proteomic analysis identified a total of 2138 proteins for both groups. There were 89 proteins found to be differentially expressed in exosomes of NSP versus SA subjects, of which 37 were increased in the NSP group and 52 were increased in the SA group. One-third of the exosomes-associated proteins highly expressed in NSP samples were involved in the reproductive process; conversely, the over-expressed proteins in exosomes of SA samples were not functionally specific. Quantitative data were confirmed on seminal exosomes from different cohorts of subjects. LARGE SCALE DATA N/A LIMITATIONS, REASONS FOR CAUTION Transfer of the proteins from exosomes to spermatozoa has been only partially demonstrated and up-take mechanisms are still poorly defined. WIDER IMPLICATIONS OF THE FINDINGS Seminal exosomes carry proteins that are potentially able to either favour or inhibit the reproductive process in humans. A better understanding of these phenomena might pave the way for novel intervention measures in terms of male infertility. STUDY FUNDING/COMPETING INTEREST(S) This study was funded by the Italian Ministry of Health through an Institution Seed Grant. None of the authors has any competing interests.

scholarly journals Alternative proteins are functional regulators in cell reprogramming by PKA activation

2020 ◽  
Vol 48 (14) ◽  
pp. 7864-7882 ◽  
Author(s):  
Tristan Cardon ◽  
Julien Franck ◽  
Etienne Coyaud ◽  
Estelle M N Laurent ◽  
Marina Damato ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Large Scale ◽  
Epithelial Mesenchymal Transition ◽  
Gene Annotation ◽  
Mass Spectrometry Analysis ◽  
Cell Reprogramming ◽  
Pathway Enrichment Analysis ◽  
Mesenchymal Transition ◽  
Spectrometry Analysis ◽  
Biological Studies

Abstract It has been recently shown that many proteins are lacking from reference databases used in mass spectrometry analysis, due to their translation templated on alternative open reading frames. This questions our current understanding of gene annotation and drastically expands the theoretical proteome complexity. The functions of these alternative proteins (AltProts) still remain largely unknown. We have developed a large-scale and unsupervised approach based on cross-linking mass spectrometry (XL-MS) followed by shotgun proteomics to gather information on the functional role of AltProts by mapping them back into known signalling pathways through the identification of their reference protein (RefProt) interactors. We have identified and profiled AltProts in a cancer cell reprogramming system: NCH82 human glioma cells after 0, 16, 24 and 48 h Forskolin stimulation. Forskolin is a protein kinase A activator inducing cell differentiation and epithelial–mesenchymal transition. Our data show that AltMAP2, AltTRNAU1AP and AltEPHA5 interactions with tropomyosin 4 are downregulated under Forskolin treatment. In a wider perspective, Gene Ontology and pathway enrichment analysis (STRING) revealed that RefProts associated with AltProts are enriched in cellular mobility and transfer RNA regulation. This study strongly suggests novel roles of AltProts in multiple essential cellular functions and supports the importance of considering them in future biological studies.

scholarly journals A holistic model for coastal flooding using system diagrams and the Source-Pathway-Receptor (SPR) concept

2012 ◽  
Vol 12 (5) ◽  
pp. 1431-1439 ◽  
Author(s):  
S. Narayan ◽  
S. Hanson ◽  
R. J. Nicholls ◽  
D. Clarke ◽  
P. Willems ◽  
...  
Keyword(s):  
Large Scale ◽  
Numerical Models ◽  
Flood Management ◽  
Coastal Flooding ◽  
System Level ◽  
Holistic Model ◽  
Directional Model ◽  
Coastal Flood ◽  
Systems Model ◽  
Level Analysis

Abstract. Coastal flooding is a problem of increasing relevance in low-lying coastal regions worldwide. In addition to the anticipated increase in likelihood and magnitude of coastal floods due to climate change, there is rapid growth in coastal assets and infrastructure. Sustainable and integrated coastal flood management over large areas and varying coastline types cannot be simply treated as local combinations of flood defences and floodplains. Rather, a system level analysis of floodplains is required to structure the problem as a first step before applying quantitative models. In this paper such a model is developed using system diagrams and the Source-Pathway-Receptor (SPR) concept, to structure our understanding of large and complex coastal flood systems. A graphical systems model is proposed for the assessment of coastal flood systems with regard to individual elements and their topological relationships. Two examples are discussed – a unidirectional model for a large-scale flood system, and a multi-directional model for a smaller-scale system, both based on the Western Scheldt estuary. The models help to develop a comprehensive understanding of system elements and their relationships and provide a holistic overview of the coastal flood system. The approach shows that a system level analysis of floodplains is more effective than simple topographic maps when conveying complex information. The models are shown to be useful as an apriori approach for making the assumptions about flood mechanisms explicit and for informing inputs to numerical models.

Integrated plant proteomics — putting the green genomes to work

2003 ◽  
Vol 30 (5) ◽  
pp. 471 ◽  
Author(s):  
Joshua L. Heazlewood ◽  
A. Harvey Millar
Keyword(s):  
Mass Spectrometry ◽  
Data Storage ◽  
Large Scale ◽  
Protein Identification ◽  
Mass Spectrometry Analysis ◽  
Plant Science ◽  
Genomic Information ◽  
Plant Proteomics ◽  
Spectrometry Analysis ◽  
Complete Set

Protein analysis has been at the heart of plant science for many years, but with new questions emerging from an abundance of genomic information and further improvements in technology, there are now new opportunities to undertake large-scale analyses and to move to more complex systems than has been possible previously. This explosion of interest and data is often referred to simply as proteomics, which is the study of the complete set of proteins expressed at a given time and place, the proteome. As its name suggests proteomics is intricately linked to allied technologies such as genomics, transcriptomics and metabolomics. In this review of plant proteomics we outline a series of issues that face the practical user, particularly the largest problem that currently faces researchers, the myriad of options to choose from. The choices, problems and pitfalls of entering into gel-based and non-gel-based arraying techniques are discussed together with advances in pre-fractionation of samples, liquid chromatography separations and subcellular analyses. Issues relating to mass spectrometry analysis and the eventual protein identification are outlined, and the dilemmas of data storage and analysis are highlighted. During this tour we provide a series of references to the literature — experimental, theoretical and technical — to illustrate the breadth of current investigations using these techniques.

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