Publications by authors named "Arthur T Kopylov"

41 Publications

Convolutional neural network in proteomics and metabolomics for determination of comorbidity between cancer and schizophrenia.

J Biomed Inform 2021 Aug 23;122:103890. Epub 2021 Aug 23.

Biobanking Group, Branch of Institute of Biomedical Chemistry "Scientific and Education Center," 10 Pogodinskaya str., 119121 Moscow, Russian Federation.

The association between cancer risk and schizophrenia is widely debated. Despite many epidemiological studies, there is still no strong evidence regarding the molecular basis for the comorbidity between these two pathological conditions. The vast majority of assays have been performed using clinical records of schizophrenic patients or those undergoing cancer treatment and monitored for sufficient time to find shared features between the considered conditions. We performed mass spectrometry-based proteomic and metabolomic investigations of patients with different cancer phenotypes (breast, ovarian, renal, and prostate) and patients with schizophrenia. The resulting vast quantity of proteomic and metabolomic data were then processed using systems biology and one-dimensional (1D) convolutional neural network (1DCNN) machine learning approaches. Traditional systematic approaches permit the segregation of schizophrenia and cancer phenotypes on the level of biological processes, while 1DCNN recognized "signatures" that could segregate distinct cancer phenotypes and schizophrenia at the comorbidity level. The designed network efficiently discriminated unrelated pathologies with a model accuracy of 0.90 and different subtypes of oncophenotypes with an accuracy of 0.94. The proposed strategy integrates systematic analysis of identified compounds and application of 1DCNN model for unidentified ones to reveal the similarity between distinct phenotypes.
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http://dx.doi.org/10.1016/j.jbi.2021.103890DOI Listing
August 2021

MAPK and Notch-Mediated Effects of Meso-Xanthin F199 Compounds on Proliferative Activity and Apoptosis of Human Melanocytes in Three-Dimensional Culture.

Biomed Res Int 2021 20;2021:8463161. Epub 2021 Jul 20.

Institute of General Pathology and Pathophysiology, 8 Baltiyskaya Str., 125315 Moscow, Russia.

Meso-Xanthin (Meso-Xanthin F199™) is a highly active antiaging injection drug of the latest generation. The main acting compound is fucoxanthin, supplemented with several growth factors, vitamins, and hyaluronic acid. Previous examination of fucoxanthin on melanocytes showed its ability to inhibit skin pigmentation through different signaling pathways focused on suppression of melanogenic-stimulating receptors. In turn, the anticancer property of fucoxanthin is realized through MAPK and PI3K pathways. We aimed to evaluate the effect of fucoxanthin and supplemented growth factors on melanocyte growth and transformation at a proteomic level. The effect of fucoxanthin on melanocytes cultivated in three-dimensional (3D) condition was examined using high-throughput proteomic and system biology approaches to disclose key molecular events of the targeted action. Our results demonstrated significant inhibition of cell differentiation and ubiquitination processes. We found that the negative regulation of and largely determines the inhibition of NF-B and MAPK2. Besides, fucoxanthin selectively inhibits cell differentiation via negative regulation of Raf signaling and the upstream activation of IL-1 signaling. It is assumed that inhibition of Raf influences the Notch-4 signaling and switches off the MAPK/MAPK2 cascade. Blockage of MAPK/MAPK2 is feasible due to suppression of Ras and NF-B by the addressed action of , , and . Suggestively, Meso-Xanthin F199™ can manage processes of proliferative activity and inhibition of apoptosis due to composition of fucoxanthin and growth-stimulating factors, which may increase the risk of skin cancer development under certain condition.
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http://dx.doi.org/10.1155/2021/8463161DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8315846PMC
September 2021

Molecular Portrait of an Athlete.

Diagnostics (Basel) 2021 Jun 15;11(6). Epub 2021 Jun 15.

Biobanking Group, Branch of Institute of Biomedical Chemistry "Scientific and Education Center", 109028 Moscow, Russia.

Sequencing of the human genome and further developments in "omics" technologies have opened up new possibilities in the study of molecular mechanisms underlying athletic performance. It is expected that molecular markers associated with the development and manifestation of physical qualities (speed, strength, endurance, agility, and flexibility) can be successfully used in the selection systems in sports. This includes the choice of sports specialization, optimization of the training process, and assessment of the current functional state of an athlete (such as overtraining). This review summarizes and analyzes the genomic, proteomic, and metabolomic studies conducted in the field of sports medicine.
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http://dx.doi.org/10.3390/diagnostics11061095DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8232626PMC
June 2021

Diversity of Plant Sterols Metabolism: The Impact on Human Health, Sport, and Accumulation of Contaminating Sterols.

Nutrients 2021 May 12;13(5). Epub 2021 May 12.

Institute of Biomedical Chemistry, Group of Biobanking, 10 Pogodinskaya Str., Bld. 8, 119121 Moscow, Russia.

The way of plant sterols transformation and their benefits for humans is still a question under the massive continuing revision. In fact, there are no receptors for binding with sterols in mammalians. However, possible biotransformation to steroids that can be catalyzed by gastro-intestinal microflora, microbial cells in prebiotics or cytochromes system were repeatedly reported. Some products of sterols metabolization are capable to imitate resident human steroids and compete with them for the binding with corresponding receptors, thus affecting endocrine balance and entire physiology condition. There are also tremendous reports about the natural origination of mammalian steroid hormones in plants and corresponding receptors for their binding. Some investigations and reports warn about anabolic effect of sterols, however, there are many researchers who are reluctant to believe in and have strong opposing arguments. We encounter plant sterols everywhere: in food, in pharmacy, in cosmetics, but still know little about their diverse properties and, hence, their exact impact on our life. Most of our knowledge is limited to their cholesterol-lowering influence and protective effect against cardiovascular disease. However, the world of plant sterols is significantly wider if we consider the thousands of publications released over the past 10 years.
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http://dx.doi.org/10.3390/nu13051623DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8150896PMC
May 2021

Mass Spectrometric Identification of Proteins Enhanced by the Atomic Force Microscopy Immobilization Surface.

Int J Mol Sci 2021 Jan 4;22(1). Epub 2021 Jan 4.

Institute of Biomedical Chemistry, 119121 Moscow, Russia.

An approach to highly-sensitive mass spectrometry detection of proteins after surface-enhanced concentrating has been elaborated. The approach is based on a combination of mass spectrometry and atomic force microscopy to detect target proteins. (1) Background: For this purpose, a technique for preliminary preparation of molecular relief surfaces formed as a result of a chemical or biospecific concentration of proteins from solution was developed and tested on several types of chip surfaces. (2) Methods: mass spectrometric identification of proteins using trailing detectors: ion trap, time of flight, orbital trap, and triple quadrupole. We used the electrospray type of ionization and matrix-assisted laser desorption/ionization. (3) Results: It is shown that when using locally functionalized atomically smooth surfaces, the sensitivity of the mass spectrometric method increases by two orders of magnitude as compared with measurements in solution. Conclusions: It has been demonstrated that the effective concentration of target proteins on specially prepared surfaces increases the concentration sensitivity of mass spectrometric detectors-time-of-flight, ion trap, triple quadrupole, and orbital ion trap in the concentration range from up to 10 M.
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http://dx.doi.org/10.3390/ijms22010431DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7795915PMC
January 2021

Pharmacogenetic Testing: A Tool for Personalized Drug Therapy Optimization.

Pharmaceutics 2020 Dec 19;12(12). Epub 2020 Dec 19.

Biobanking Group, Branch of Institute of Biomedical Chemistry "Scientific and Education Center", 109028 Moscow, Russia.

Pharmacogenomics is a study of how the genome background is associated with drug resistance and how therapy strategy can be modified for a certain person to achieve benefit. The pharmacogenomics (PGx) testing becomes of great opportunity for physicians to make the proper decision regarding each non-trivial patient that does not respond to therapy. Although pharmacogenomics has become of growing interest to the healthcare market during the past five to ten years the exact mechanisms linking the genetic polymorphisms and observable responses to drug therapy are not always clear. Therefore, the success of PGx testing depends on the physician's ability to understand the obtained results in a standardized way for each particular patient. The review aims to lead the reader through the general conception of PGx and related issues of PGx testing efficiency, personal data security, and health safety at a current clinical level.
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http://dx.doi.org/10.3390/pharmaceutics12121240DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7765968PMC
December 2020

Molecular pathophysiology of diabetes mellitus during pregnancy with antenatal complications.

Sci Rep 2020 11 12;10(1):19641. Epub 2020 Nov 12.

Department of Pathology, Institute of General Pathology and Pathophysiology, 8 Baltyiskaya str., 125315, Moscow, Russia.

Gestational diabetes mellitus is a daunting problem accompanied by severe fetal development complications and type 2 diabetes mellitus in postpartum. Diagnosis of diabetic conditions occurs only in the second trimester, while associated antenatal complications are typically revealed even later. We acquired an assay of peripheral and cord blood samples of patients with different types of diabetes mellitus who delivered either healthy newborns or associated with fetopathy complications. Obtained data were handled with qualitative and quantitative analysis. Pathways of molecular events involved in diabetes mellitus and fetopathy were reconstructed based on the discovered markers and their quantitative alteration. Plenty of pathways were integrated to differentiate the type of diabetes and to recognize the impact of the diabetic condition on fetal development. The impaired triglycerides transport, glucose uptake, and consequent insulin resistance are mostly affected by faulted lipid metabolism (APOM, APOD, APOH, APOC1) and encouraged by oxidative stress (CP, TF, ORM2) and inflammation (CFH, CFB, CLU) as a secondary response accompanied by changes in matrix architecture (AFM, FBLN1, AMBP). Alterations in proteomes of peripheral and cord blood were expectedly unequal. Both up- and downregulated markers were accommodated in the cast of molecular events interconnected with the lipid metabolism, RXR/PPAR-signaling pathway, and extracellular architecture modulation. The obtained results congregate numerous biological processes to molecular events that underline diabetes during gestation and uncover some critical aspects affecting fetal growth and development.
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http://dx.doi.org/10.1038/s41598-020-76689-9DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7665025PMC
November 2020

Association of Proteins Modulating Immune Response and Insulin Clearance During Gestation with Antenatal Complications in Patients with Gestational or Type 2 Diabetes Mellitus.

Cells 2020 04 21;9(4). Epub 2020 Apr 21.

Department of Pathology, Institute of General Pathology and Pathophysiology, 125315 Moscow, Russia.

Background: The purpose of the study is to establish and quantitatively assess protein markers and their combination in association with insulin uptake that may be have value for early prospective recognition of diabetic fetopathy (DF) as a complication in patients with diabetes mellitus during gestation.

Methods: Proteomic surveying and accurate quantitative measurement of selected proteins from plasma samples collected from the patients with gestational diabetes mellitus (GDM) and type 2 diabetes mellitus (T2DM) who gave birth of either healthy or affected by maternal diabetes newborns was performed using mass spectrometry.

Results: We determined and quantitatively measured several proteins, including CRP, CEACAM1, CNDP1 and Ig-family that were significantly differed in patients that gave birth of newborns with signs of DF. We found that patients with newborns associated with DF are characterized by significantly decreased CEACAM1 (113.18 ± 16.23 ng/mL and 81.09 ± 10.54 ng/mL in GDM and T2DM, < 0.005) in contrast to control group (515.6 ± 72.14 ng/mL, < 0.005). On the contrary, the concentration of CNDP1 was increased in DF-associated groups and attained 49.3 ± 5.18 ng/mL and 37.7 ± 3.34 ng/mL ( < 0.005) in GDM and T2DM groups, respectively. Among other proteins, dramatically decreased concentration of IgG4 and IgA2 subclasses of immunoglobulins were noticed.

Conclusion: The combination of the measured markers may assist (AUC = 0.893 (CI 95%, 0.785-0.980) in establishing the clinical finding of the developing DF especially in patients with GDM who are at the highest risk of chronic insulin resistance.
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http://dx.doi.org/10.3390/cells9041032DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7226479PMC
April 2020

Revelation of Proteomic Indicators for Colorectal Cancer in Initial Stages of Development.

Molecules 2020 Jan 31;25(3). Epub 2020 Jan 31.

V.N. Orekhovich Institute of Biomedical Chemistry, 119121 Moscow, Russia.

Colorectal cancer (CRC) at a current clinical level is still hardly diagnosed, especially with regard to nascent tumors, which are typically asymptotic. Searching for reliable biomarkers of early diagnosis is an extremely essential task. Identification of specific post-translational modifications (PTM) may also significantly improve net benefits and tailor the process of CRC recognition. We examined depleted plasma samples obtained from 41 healthy volunteers and 28 patients with CRC at different stages to conduct comparative proteome-scaled analysis. The main goal of the study was to establish a constellation of protein markers in combination with their PTMs and semi-quantitative ratios that may support and realize the distinction of CRC until the disease has a poor clinical manifestation. Proteomic analysis revealed 119 and 166 proteins for patients in stages I-II and III-IV, correspondingly. Plenty of proteins (44 proteins) reflected conditions of the immune response, lipid metabolism, and response to stress, but only a small portion of them were significant ( < 0.01) for distinguishing stages I-II of CRC. Among them, some cytokines (Clusterin (CLU), C4b-binding protein (C4BP), and CD59 glycoprotein (CD59), etc.) were the most prominent and the lectin pathway was specifically enhanced in patients with CRC. Significant alterations in Inter-alpha-trypsin inhibitor heavy chains (ITIH1, ITIH2, ITIH3, and ITIH4) levels were also observed due to their implication in tumor growth and the malignancy process. Other markers (Alpha-1-acid glycoprotein 2 (ORM2), Alpha-1B-glycoprotein (A1BG), Haptoglobin (HP), and Leucine-rich alpha-2-glycoprotein (LRG1), etc.) were found to create an ambiguous core involved in cancer development but also to exactly promote tumor progression in the early stages. Additionally, we identified post-translational modifications, which according to the literature are associated with the development of colorectal cancer, including kininogen 1 protein (T327-p), alpha-2-HS-glycoprotein (S138-p) and newly identified PTMs, i.e., vitamin D-binding protein (K75-ac and K370-ac) and plasma protease C1 inhibitor (Y294-p), which may also contribute and negatively impact on CRC progression. The contribution of cytokines and proteins of the extracellular matrix is the most significant factor in CRC development in the early stages. This can be concluded since tumor growth is tightly associated with chronic aseptic inflammation and concatenated malignancy related to loss of extracellular matrix stability. Due attention should be paid to Apolipoprotein E (APOE), Apolipoprotein C1 (APOC1), and Apolipoprotein B-100 (APOB) because of their impact on the malfunction of DNA repair and their capability to regulate mTOR and PI3K pathways. The contribution of the observed PTMs is still equivocal, but a significant decrease in the likelihood between modified and native proteins was not detected confidently.
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http://dx.doi.org/10.3390/molecules25030619DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7036866PMC
January 2020

Assessment of Serological Early Biomarker Candidates for Lung Adenocarcinoma by using Multiple Reaction Monitoring-Mass Spectrometry.

Proteomics Clin Appl 2020 07 16;14(4):e1900095. Epub 2020 Feb 16.

Department of Environmental and Occupational Health, College of Medicine, National Cheng Kung University, Tainan, 701, Taiwan.

Purpose: Plasma markers that enable diagnosis in the early stage of lung cancer is not discovered. A liquid chromatography multiple reaction monitoring-mass spectrometry (LC-MRM-MS) assay for identifying potential early marker proteins for lung adenocarcinoma is developed.

Experimental Design: LC-MRM-MS assay is used for measuring the level of 35 candidate peptides in plasma from 102 lung adenocarcinoma patients (including n = 50, 16, 24, and 12 in stage I, II, III, and IV, respectively.) and 84 healthy controls. Stable isotope labeled standard peptides are synthesized to accurately measure the amount of these proteins.

Results: Seven proteins are able to distinguish stage I patients from controls. These proteins are combined in to a protein marker panel which improve the sensitivity to discriminate stage I patients from controls with cross-validated area under the curve = 0.76. Besides, it is found that low expression of eukaryotic initiation factor 4A-I and high expression of lumican show significantly poor prognosis in overall survival (p = 0.012 and 0.0074, respectively), which may be used as prognostic biomarkers for lung cancer.

Conclusions And Clinical Relevance: Proteins highlighted here may be used for early detection of lung adenocarcinoma or therapeutics development after validation in a larger cohort.
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http://dx.doi.org/10.1002/prca.201900095DOI Listing
July 2020

Challenges of the Human Proteome Project: 10-Year Experience of the Russian Consortium.

J Proteome Res 2019 12 28;18(12):4206-4214. Epub 2019 Oct 28.

Institute of Biomedical Chemistry , Moscow 119435 , Russia.

This manuscript collects all the efforts of the Russian Consortium, bottlenecks revealed in the course of the C-HPP realization, and ways of their overcoming. One of the main bottlenecks in the C-HPP is the insufficient sensitivity of proteomic technologies, hampering the detection of low- and ultralow-copy number proteins forming the "dark part" of the human proteome. In the frame of MP-Challenge, to increase proteome coverage we suggest an experimental workflow based on a combination of shotgun technology and selected reaction monitoring with two-dimensional alkaline fractionation. Further, to detect proteins that cannot be identified by such technologies, nanotechnologies such as combined atomic force microscopy with molecular fishing and/or nanowire detection may be useful. These technologies provide a powerful tool for single molecule analysis, by analogy with nanopore sequencing during genome analysis. To systematically analyze the functional features of some proteins (CP50 Challenge), we created a mathematical model that predicts the number of proteins differing in amino acid sequence: proteoforms. According to our data, we should expect about 100 000 different proteoforms in the liver tissue and a little more in the HepG2 cell line. The variety of proteins forming the whole human proteome significantly exceeds these results due to post-translational modifications (PTMs). As PTMs determine the functional specificity of the protein, we propose using a combination of gene-centric transcriptome-proteomic analysis with preliminary fractionation by two-dimensional electrophoresis to identify chemically modified proteoforms. Despite the complexity of the proposed solutions, such integrative approaches could be fruitful for MP50 and CP50 Challenges in the framework of the C-HPP.
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http://dx.doi.org/10.1021/acs.jproteome.9b00358DOI Listing
December 2019

Multi-dimensional immunoproteomics coupled with in vitro recapitulation of oncogenic NRAS identifies diagnostically relevant autoantibody biomarkers in thyroid neoplasia.

Cancer Lett 2019 12 18;467:96-106. Epub 2019 Jul 18.

Engelhardt Institute of Molecular Biology, Russian Academy of Sciences, Moscow, Russia.

Tumor-associated antigen (TAA)-specific autoantibodies have been widely implicated in cancer diagnosis. However, cancer cell lines that are typically exploited as candidate TAA sources in immunoproteomic studies may fail to accurately represent the autoantigen-ome of lower-grade neoplasms. Here, we established an integrated strategy for the identification of disease-relevant TAAs in thyroid neoplasia, which combined NRAS oncogene expression in non-tumorous thyroid Nthy-ori 3-1 cells with a multi-dimensional proteomic technique DISER that consisted of profiling NRAS-induced proteins using 2-dimensional difference gel electrophoresis (2D-DIGE) coupled with serological proteome analysis (SERPA) of the TAA repertoire of patients with thyroid encapsulated follicular-patterned/RAS-like phenotype (EFP/RLP) tumors. We identified several candidate cell-based (nicotinamide phosphoribosyltransferase NAMPT, glutamate dehydrogenase GLUD1, and glutathione S-transferase omega-1 GSTO1) and autoantibody (fumarate hydratase FH, calponin-3 CNN3, and pyruvate kinase PKM autoantibodies) biomarkers, including NRAS-induced TAA phosphoglycerate kinase 1 PGK1. Meta-profiling of the reactivity of the identified autoantibodies across an independent SERPA series implicated the PKM autoantibody as a histological phenotype-independent biomarker of thyroid malignancy (11/38 (29%) patients with overtly malignant and uncertain malignant potential (UMP) tumors vs 0/22 (p = 0.0046) and 0/20 (p = 0.011) patients with non-invasive EFP/RLP tumors and healthy controls, respectively). PGK1 and CNN3 autoantibodies were identified as EFP/RLP-specific biomarkers, potentially suitable for further discriminating tumors with different malignant potential (PGK1: 7/22 (32%) patients with non-invasive EFP/RLP tumors vs 0/38 (p = 0.00044) and 0/20 (p = 0.0092) patients with other tumors and healthy controls, respectively; СNN3: 9/29 (31%) patients with malignant and borderline EFP/RLP tumors vs 0/31 (p = 0.00068) and 0/20 (p = 0.0067) patients with other tumors and healthy controls, respectively). The combined use of PKM, CNN3, and PGK1 autoantibodies allowed the reclassification of malignant/UMP tumor risk in 19/41 (46%) of EFP/RLP tumor patients. Taken together, we established an experimental pipeline DISER for the concurrent identification of cell-based and TAA biomarkers. The combination of DISER with in vitro oncogene expression allows further targeted identification of oncogene-induced TAAs. Using this integrated approach, we identified candidate autoantibody biomarkers that might be of value for differential diagnostic purposes in thyroid neoplasia.
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http://dx.doi.org/10.1016/j.canlet.2019.07.013DOI Listing
December 2019

Affinity Isolation and Mass Spectrometry Identification of Prostacyclin Synthase (PTGIS) Subinteractome.

Biology (Basel) 2019 Jun 20;8(2). Epub 2019 Jun 20.

Department of Proteomic Research and Mass Spectrometry, Institute of Biomedical Chemistry (IBMC), 10 Pogodinskaya str., 119121 Moscow, Russia.

Prostacyclin synthase (PTGIS; EC 5.3.99.4) catalyzes isomerization of prostaglandin H to prostacyclin, a potent vasodilator and inhibitor of platelet aggregation. At present, limited data exist on functional coupling and possible ways of regulating PTGIS due to insufficient information about protein-protein interactions in which this crucial enzyme is involved. The aim of this study is to isolate protein partners for PTGIS from rat tissue lysates. Using CNBr-activated Sepharose 4B with covalently immobilized PTGIS as an affinity sorbent, we confidently identified 58 unique proteins by mass spectrometry (LC-MS/MS). The participation of these proteins in lysate complex formation was characterized by SEC lysate profiling. Several potential members of the PTGIS subinteractome have been validated by surface plasmon resonance (SPR) analysis. SPR revealed that PTGIS interacted with full-length cytochrome P450 2J2 and glutathione S-transferase (GST). In addition, PTGIS was shown to bind synthetic peptides corresponding to sequences of for GSTA1, GSTM1, aldo-keto reductase (AKR1A1), glutaredoxin 3 (GLRX3) and histidine triad nucleotide binding protein 2 (HINT2). Prostacyclin synthase could potentially be involved in functional interactions with identified novel protein partners participating in iron and heme metabolism, oxidative stress, xenobiotic and drugs metabolism, glutathione and prostaglandin metabolism. The possible biological role of the recognized interaction is discussed in the context of PTGIS functioning.
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http://dx.doi.org/10.3390/biology8020049DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6628129PMC
June 2019

200+ Protein Concentrations in Healthy Human Blood Plasma: Targeted Quantitative SRM SIS Screening of Chromosomes 18, 13, Y, and the Mitochondrial Chromosome Encoded Proteome.

J Proteome Res 2019 01 11;18(1):120-129. Epub 2018 Dec 11.

Institute of Biomedical Chemistry , Moscow 119435 , Russia.

This work continues the series of the quantitative measurements of the proteins encoded by different chromosomes in the blood plasma of a healthy person. Selected Reaction Monitoring with Stable Isotope-labeled peptide Standards (SRM SIS) and a gene-centric approach, which is the basis for the implementation of the international Chromosome-centric Human Proteome Project (C-HPP), were applied for the quantitative measurement of proteins in human blood plasma. Analyses were carried out in the frame of C-HPP for each protein-coding gene of the four human chromosomes: 18, 13, Y, and mitochondrial. Concentrations of proteins encoded by 667 genes were measured in 54 blood plasma samples of the volunteers, whose health conditions were consistent with requirements for astronauts. The gene list included 276, 329, 47, and 15 genes of chromosomes 18, 13, Y, and the mitochondrial chromosome, respectively. This paper does not make claims about the detection of missing proteins. Only 205 proteins (30.7%) were detected in the samples. Of them, 84, 106, 10, and 5 belonged to chromosomes 18, 13, and Y and the mitochondrial chromosome, respectively. Each detected protein was found in at least one of the samples analyzed. The SRM SIS raw data are available in the ProteomeXchange repository (PXD004374, PASS01192).
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http://dx.doi.org/10.1021/acs.jproteome.8b00391DOI Listing
January 2019

Proteomic profiling data of HEK293 proteins bound to human recombinant renalases-1 and -2.

Data Brief 2018 Dec 30;21:1477-1482. Epub 2018 Oct 30.

Institute of Biomedical Chemistry, 10 Pogodinskaya street, Moscow 119121, Russia.

Renalase (RNLS) is a recently discovered protein involved in blood pressure regulation. It exists both as an intracellular catalytically active flavoprotein (EC 1.6.3.5 dihydro-NAD(P):oxygen oxidoreductase) and an extracellular protein that demonstrates various cell protecting effects. Using a twenty-membered peptide corresponding to the residues 220-239 of the renalase sequence (RP-220) and the HK-2 cell line Wang et al. identified a renalase-binding protein, which was considered as a receptor for extracellular renalase crucial for MAPK signaling (Wang et al., 2015) [1]. In this study we have investigated profiles of renalase binding proteins in HEK293 cells by using affinity based proteomic profiling with full-length recombinant human RNLS-1 and human RNLS-2 as affinity ligands followed by analysis of bound proteins by liquid chromatography-mass spectrometry. Both renalases (RNLS-1 and RNLS-2) contain the RP-220 sequence (residues 220-239) but differ in their C-terminal region (residues 293-342 and 293-325, respectively). Profiling of HEK293 proteins resulted in identification of two different sets of proteins specifically bound to RNLS-1 and RNLS-2, respectively. We thus demonstrate that the C-terminal region is crucial for specific binding of renalase to its targets and/or receptors.
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http://dx.doi.org/10.1016/j.dib.2018.10.137DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6234383PMC
December 2018

Increased Sensitivity of Mass Spectrometry by Alkaline Two-Dimensional Liquid Chromatography: Deep Cover of the Human Proteome in Gene-Centric Mode.

J Proteome Res 2018 12 19;17(12):4258-4266. Epub 2018 Nov 19.

Institute of Biomedical Chemistry, RAS , Moscow , Russia.

Currently, great interest is paid to the identification of "missing" proteins that have not been detected in any biological material at the protein level (PE1). In this paper, using the Universal Proteomic Standard sets 1 and 2 (UPS1 and UPS2, respectively) as an example, we characterized mass spectrometric approaches from the point of view of sensitivity (Sn), specificity (Sp), and accuracy (Ac). The aim of the paper was to show the utility of a mass spectra approach for protein detection. This sets consists of 48 high-purity human proteins without single aminoacid polymorphism (SAP) or post translational modification (PTM). The UPS1 set consists of the same 48 proteins at 5 pmols each, and in UPS2, proteins were grouped into 5 groups in accordance with their molar concentration, ranging from 10 to 10 M. Single peptides from the 92% and 96% of all sets of proteins could be detected in a pure solution of UPS2 and UPS1, respectively, by selected reaction monitoring with stable isotope-labeled standards (SRM-SIS). We also found that, in the presence of a biological matrix such as Escherichia coli extract or human blood plasma (HBP), SRM-SIS makes it possible to detect from 63% to 79% of proteins in the UPS2 set (sensitivity) with the highest specificity (∼100%) and an accuracy of 80% by increasing the sensitivity of shotgun and selected reaction monitoring combined with a stable-isotope-labeled peptide standard (SRM-SIS technology) by fractionating samples using reverse-phase liquid chromatography under alkaline conditions (2D-LC_alk). It is shown that this technique of sample fractionation allows the SRM-SIS to detect 98% of the single peptides from the proteins present in the pure solution of UPS2 (47 out of 48 proteins). When the extracts of E. coli or Pichia pastoris are added as biological matrixes to the UPS2, 46, and 45 out of 48 proteins (∼95%) can be detected, respectively, using the SRM-SIS combined with 2D-LC_alk. The combination of the 2D-LC_alk SRM-SIS and shotgun technologies allows us to increase the sensitivity up to 100% in the case of the proteins of the UPS2 set. The usage of that technology can be a solution for identifying the so-called "missing" proteins and, eventually, creating the deep proteome of a particular chromosome of tissue or organs. Experimental data have been deposited in the PeptideAtlas SRM Experiment Library with the dataset identifier PASS01192 and the PRIDE repository with the dataset identifier PXD007643.
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http://dx.doi.org/10.1021/acs.jproteome.8b00754DOI Listing
December 2018

Neurotoxic Effects of Aβ6-42 Peptides Mimicking Putative Products Formed by the Angiotensin Converting Enzyme.

J Alzheimers Dis 2018 ;66(1):263-270

Engelhardt Institute of Molecular Biology, Russian Academy of Sciences, Moscow, Russia.

Angiotensin converting enzyme (ACE) is involved in proteolytic processing of the amyloid-β(Aβ) peptide implicated in the development of Alzheimer's disease (AD) and known products of ACE-based processing of Aβ42 are characterized by reduced aggregability and cytotoxicity. Recently it has been demonstrated that ACE can act as an arginine specific endopeptidase cleaving the N-terminal pentapeptide (Aβ1-5) from synthetic Aβ peptide analogues. In the context of proteolytic processing of full length Aβ42, this suggests possible formation of Aβ6-42 species. The aim of this study was to test a hypothesis that some N-terminally truncated Aβ peptide(s) could retain aggregability and neurotoxic properties typical for Aβ42. We have investigated aggregability of two amyloid-β peptides, Aβ6-42 and isoD7-Aβ6-42, mimicking potential proteolytic products of Aβ42 and isoD7-Aβ42, and evaluated their effects on the repertoire of brain Aβ binding proteins, and cytotoxicity towards neuroblastoma SH-SY5Y cells. Aggregability of isoD7-Aβ6-42 and Aβ6-42 was higher than that of full-length peptides Aβ42 and isoD7-Aβ42, while the repertoire of mouse brain Aβ binding proteins dramatically decreased. Aβ6-42 and isoD7-Aβ6-42 exhibited higher neurotoxicity towards SH-SY5Y cells than Aβ42 and isoD7-Aβ42, respectively. They effectively stimulated production of ROS and NO, and also TNFα secretion by cells. Thus, our results suggest that ACE-dependent processing of full-length Aβs could result in formation of more pathogenic peptides.
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http://dx.doi.org/10.3233/JAD-180500DOI Listing
September 2019

Next Steps on in Silico 2DE Analyses of Chromosome 18 Proteoforms.

J Proteome Res 2018 12 2;17(12):4085-4096. Epub 2018 Oct 2.

Institute of Biomedical Chemistry of Russian Academy of Medical Sciences , Pogodinskaya 10 , Moscow 119121 , Russia.

In the boundaries of the chromosome-centric Human Proteome Project (c-HPP) to obtain information about proteoforms coded by chromosome 18, several cell lines (HepG2, glioblastoma, LEH), normal liver, and plasma were analyzed. In our study, we have been using proteoform separation by two-dimensional electrophoresis (2DE) (a sectional analysis) and a semivirtual 2DE with following shotgun mass spectrometry using LC-ESI-MS/MS. Previously, we published a first draft of this research, where only HepG2 cells were tested. Here, we present the next step using more detailed analysis and more samples. Altogether, confident (2 significant sequences minimum) information about proteoforms of 117 isoforms coded by 104 genes of chromosome 18 was obtained. The 3D-graphs showing distribution of different proteoforms from the same gene in the 2D map were generated. Additionally, a semivirtual 2DE approach has allowed for detecting more proteoforms and estimating their pI more precisely. Data are available via ProteomeXchange with identifier PXD010142.
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http://dx.doi.org/10.1021/acs.jproteome.8b00386DOI Listing
December 2018

FractionOptimizer: a method for optimal peptide fractionation in bottom-up proteomics.

Anal Bioanal Chem 2018 Jun 17;410(16):3827-3833. Epub 2018 Apr 17.

V.L. Talrose Institute for Energy Problems of Chemical Physics, RAS, Moscow, 119334, Russia.

Recent advances in mass spectrometry and separation technologies created the opportunities for deep proteome characterization using shotgun proteomics approaches. The "real world" sample complexity and high concentration range limit the sensitivity of this characterization. The common strategy for increasing the sensitivity is sample fractionation prior to analysis either at the protein or the peptide level. Typically, fractionation at the peptide level is performed using linear gradient high-performance liquid chromatography followed by uniform fraction collection. However, this way of peptide fractionation results in significantly suboptimal operation of the mass spectrometer due to the non-uniform distribution of peptides between the fractions. In this work, we propose an approach based on peptide retention time prediction allowing optimization of chromatographic conditions and fraction collection procedures. An open-source software implementing the approach called FractionOptimizer was developed and is available at http://hg.theorchromo.ru/FractionOptimizer . The performance of the developed tool was demonstrated for human embryonic kidney (HEK293) cell line lysate. In these experiments, we improved the uniformity of the peptides distribution between fractions. Moreover, in addition to 13,492 peptides, we found 6787 new peptides not identified in the experiments without fractionation and up to 800 new proteins (or 25%). Graphical abstract The analysis workflow employing FractionOptimizer software.
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http://dx.doi.org/10.1007/s00216-018-1054-2DOI Listing
June 2018

Brochosomins and other novel proteins from brochosomes of leafhoppers (Insecta, Hemiptera, Cicadellidae).

Insect Biochem Mol Biol 2018 03 10;94:10-17. Epub 2018 Jan 10.

Department of Entomology, University of Illinois at Urbana-Champaign, Urbana, 61801, IL, USA. Electronic address:

Brochosomes (BS) are secretory granules resembling buckyballs, produced intracellularly in specialized glandular segments of the Malpighian tubules and forming superhydrophobic coatings on the integuments of leafhoppers (Hemiptera, Cicadellidae). Their composition is poorly known. Using a combination of SDS-PAGE, LC-MS/MS, next-generation sequencing (RNAseq) and bioinformatics we demonstrate that the major structural component of BS of the leafhopper Graphocephala fennahi Young is a novel family of 21-40-kDa secretory proteins, referred to herein as brochosomins (BSM), apparently cross-linked by disulfide bonds. At least 28 paralogous BSM were identified in a transcriptome assembly of this species, most of which were detected in BS. Multiple additional BS-associated proteins (BSAP), possibly loosely attached to the outer and inner surfaces of BS, were also identified; some of these were glycine-, tyrosine- and proline-rich. BSM and BSAP together accounted for half of the 100 most expressed transcripts in the Malpighian tubules of G. fennahi. Except for several minor BSAP possibly related to cyclases, BSM and BSAP had no homologs among known proteins, thus representing taxonomically restricted gene families (orphans). Searching in 50 whole-body transcriptome assemblies of Hemiptera found homologs of BSM in representatives of all five families of the superfamily Membracoidea (Cicadellidae, Myerslopiidae, Aetalionidae, Membracidae, and Melizoderidae), but not in other lineages. Among the identified proteins only BSM were shared in common between all 17 surveyed leafhoppers known to produce BS. Combined CHN elemental and aminoacid analyses estimated the total protein content of BS from the integument of G. fennahi to be 60-70%.
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http://dx.doi.org/10.1016/j.ibmb.2018.01.001DOI Listing
March 2018

Quantitative assessment of betamethasone dual-acting formulation in urine of patients with rheumatoid arthritis and ankylosing spondylitis after single-dose intramuscular administration and its application to long-term pharmacokinetic study.

J Pharm Biomed Anal 2018 Feb 6;149:278-289. Epub 2017 Nov 6.

V. N. Orekhovich Research Institute of Biomedical Chemistry, 10 Pogodinskaya str. bld. 8, 119121 Moscow, Russian Federation.

Quantitative evaluation and assessment of pharmacokinetic parameters of Diprospan (suspension for injection 7mg/mL (2mg+5mg/mL) of betamethasone) were performed in urine samples taken from patients with rheumatoid arthritis or ankylosing spondylitis for 28days after systemic intramuscular administration in routine clinical practice in an open-comparative prospective cohort study. The maximum betamethasone concentration was reached at day 4 of the follow-up; in some cases, β-phase of elimination of the drug was appeared at day 14 or at day 21 of the follow-up. The deferred β-phase elimination was likely a consequence of the physiological characteristics of the patients or of the influence of non-steroidal agents. The half-life of betamethasone was 8.5days. The elimination rate constant was 2.49h-1; the mean clearance was 4.72L/d. The recommended frequency of the drug administration to its complete elimination was estimated up to 48days. Mann-Whitney test showed no significant differences in pharmacokinetic characteristics between male and female subjects. The prolonged elimination phase was observed in patients with deviations in their body mass index, continual treatment by diclofenac and nimesulide or, possibly, after consuming an alcohol. The study was recorded in Clinical Trials open source with identifier NCT03119454.
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http://dx.doi.org/10.1016/j.jpba.2017.11.021DOI Listing
February 2018

Why Are the Correlations between mRNA and Protein Levels so Low among the 275 Predicted Protein-Coding Genes on Human Chromosome 18?

J Proteome Res 2017 12 27;16(12):4311-4318. Epub 2017 Oct 27.

Institute of Biomedical Chemistry RAS , 119121 Moscow, Russia.

In this work targeted (selected reaction monitoring, SRM, PASSEL: PASS00697) and panoramic (shotgun LC-MS/MS, PRIDE: PXD00244) mass-spectrometric methods as well as transcriptomic analysis of the same samples using RNA-Seq and PCR methods (SRA experiment IDs: SRX341198, SRX267708, SRX395473, SRX390071) were applied for quantification of chromosome 18 encoded transcripts and proteins in human liver and HepG2 cells. The obtained data was used for the estimation of quantitative mRNA-protein ratios for the 275 genes of the selected chromosome in the selected tissues. The impact of methodological limitations of existing analytical proteomic methods on gene-specific mRNA-protein ratios and possible ways of overcoming these limitations for detection of missing proteins are also discussed.
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http://dx.doi.org/10.1021/acs.jproteome.7b00348DOI Listing
December 2017

Constitutive and activation-dependent phosphorylation of lymphocyte phosphatase-associated phosphoprotein (LPAP).

PLoS One 2017 21;12(8):e0182468. Epub 2017 Aug 21.

NRC Institute of Immunology FMBA of Russia, Moscow, Russia.

Lymphocyte phosphatase-associated phosphoprotein (LPAP) is a small transmembrane protein expressed exclusively in lymphocytes. LPAP is a component of a supramolecular complex composed of the phosphatase CD45, the co-receptor CD4, and the kinase Lck. In contrast to its immunologically important partners, the function of LPAP is unknown. We hypothesized that the biological role of LPAP may be determined by analyzing LPAP phosphorylation. In the present study, we identified LPAP phosphorylation sites by site-directed mutagenesis, phospho-specific antibodies, and protein immunoprecipitation coupled to mass spectrometry analysis. Our results confirmed previous reports that Ser-99, Ser-153, and Ser-163 are phosphorylated, as well as provided evidence for the phosphorylation of Ser-172. Using various SDS-PAGE techniques, we detected and quantified a set of LPAP phosphoforms that were assigned to a combination of particular phosphorylation events. The phosphorylation of LPAP appears to be a tightly regulated process. Our results support the model: following phorbol 12-myristate 13-acetate (PMA) or TCR/CD3 activation of T cells, LPAP is rapidly dephosphorylated at Ser-99 and Ser-172 and slowly phosphorylated at Ser-163. Ser-153 exhibited a high basal level of phosphorylation in both resting and activated cells. Therefore, we suggest that LPAP may function as a co-regulator of T-cell signaling.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0182468PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5565103PMC
October 2017

Highly sensitive protein detection by biospecific AFM-based fishing with pulsed electrical stimulation.

FEBS Open Bio 2017 08 10;7(8):1186-1195. Epub 2017 Jul 10.

Institute of Biomedical Chemistry Moscow Russia.

We report here the highly sensitive detection of protein in solution at concentrations from 10 to 10 m using the combination of atomic force microscopy (AFM) and mass spectrometry. Biospecific detection of biotinylated bovine serum albumin was carried out by fishing out the protein onto the surface of AFM chips with immobilized avidin, which determined the specificity of the analysis. Electrical stimulation was applied to enhance the fishing efficiency. A high sensitivity of detection was achieved by application of nanosecond electric pulses to highly oriented pyrolytic graphite placed under the AFM chip. A peristaltic pump-based flow system, which is widely used in routine bioanalytical assays, was employed throughout the analysis. These results hold promise for the development of highly sensitive protein detection methods using nanosensor devices.
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http://dx.doi.org/10.1002/2211-5463.12253DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5537060PMC
August 2017

A multicentric study to evaluate the use of relative retention times in targeted proteomics.

J Proteomics 2017 01 29;152:138-149. Epub 2016 Oct 29.

ProteoRed-ISCIII, Centro Nacional de Investigaciones Cardiovasculares Carlos III (CNIC), Madrid 28029, Spain.

Despite the maturity reached by targeted proteomic strategies, reliable and standardized protocols are urgently needed to enhance reproducibility among different laboratories and analytical platforms, facilitating a more widespread use in biomedical research. To achieve this goal, the use of dimensionless relative retention times (iRT), defined on the basis of peptide standard retention times (RT), has lately emerged as a powerful tool. The robustness, reproducibility and utility of this strategy were examined for the first time in a multicentric setting, involving 28 laboratories that included 24 of the Spanish network of proteomics laboratories (ProteoRed-ISCIII). According to the results obtained in this study, dimensionless retention time values (iRTs) demonstrated to be a useful tool for transferring and sharing peptide retention times across different chromatographic set-ups both intra- and inter-laboratories. iRT values also showed very low variability over long time periods. Furthermore, parallel quantitative analyses showed a high reproducibility despite the variety of experimental strategies used, either MRM (multiple reaction monitoring) or pseudoMRM, and the diversity of analytical platforms employed.

Biological Significance: From the very beginning of proteomics as an analytical science there has been a growing interest in developing standardized methods and experimental procedures in order to ensure the highest quality and reproducibility of the results. In this regard, the recent (2012) introduction of the dimensionless retention time concept has been a significant advance. In our multicentric (28 laboratories) study we explore the usefulness of this concept in the context of a targeted proteomics experiment, demonstrating that dimensionless retention time values is a useful tool for transferring and sharing peptide retention times across different chromatographic set-ups.
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http://dx.doi.org/10.1016/j.jprot.2016.10.014DOI Listing
January 2017

State of the Art of Chromosome 18-Centric HPP in 2016: Transcriptome and Proteome Profiling of Liver Tissue and HepG2 Cells.

J Proteome Res 2016 11 29;15(11):4030-4038. Epub 2016 Aug 29.

Institute of Biomedical Chemistry , Pogodinskaya Street, 10, Moscow 119121, Russia.

A gene-centric approach was applied for a large-scale study of expression products of a single chromosome. Transcriptome profiling of liver tissue and HepG2 cell line was independently performed using two RNA-Seq platforms (SOLiD and Illumina) and also by Droplet Digital PCR (ddPCR) and quantitative RT-PCR. Proteome profiling was performed using shotgun LC-MS/MS as well as selected reaction monitoring with stable isotope-labeled standards (SRM/SIS) for liver tissue and HepG2 cells. On the basis of SRM/SIS measurements, protein copy numbers were estimated for the Chromosome 18 (Chr 18) encoded proteins in the selected types of biological material. These values were compared with expression levels of corresponding mRNA. As a result, we obtained information about 158 and 142 transcripts for HepG2 cell line and liver tissue, respectively. SRM/SIS measurements and shotgun LC-MS/MS allowed us to detect 91 Chr 18-encoded proteins in total, while an intersection between the HepG2 cell line and liver tissue proteomes was ∼66%. In total, there were 16 proteins specifically observed in HepG2 cell line, while 15 proteins were found solely in the liver tissue. Comparison between proteome and transcriptome revealed a poor correlation (R ≈ 0.1) between corresponding mRNA and protein expression levels. The SRM and shotgun data sets (obtained during 2015-2016) are available in PASSEL (PASS00697) and ProteomeExchange/PRIDE (PXD004407). All measurements were also uploaded into the in-house Chr 18 Knowledgebase at http://kb18.ru/protein/matrix/416126 .
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http://dx.doi.org/10.1021/acs.jproteome.6b00380DOI Listing
November 2016

Dataset of target mass spectromic proteome profiling for human chromosome 18.

Data Brief 2016 Sep 26;8:1365-9. Epub 2016 Jul 26.

Orekhovich Institute of Biomedical Chemistry, Moscow, Russia.

Proteome profiling is a type of quantitative analysis that reveals level of protein expression in the sample. Proteome profiling by using selected reaction monitoring is an approach for the Chromosome-centric Human Proteome Project (C-HPP). Here we describe dataset generated in the course of the pilot phase of Russian part of C-HPP, which was focused on human Chr 18 proteins. Proteome profiling was performed using stable isotope-labeled standards (SRM/SIS) for plasma, liver tissue and HepG2 cells. Dataset includes both positive and negative results of protein detection. These data were partly discussed in recent publications, "Chromosome 18 Transcriptome Profiling and Targeted Proteome Mapping in Depleted Plasma, Liver Tissue and HepG2 Cells" [1] and "Chromosome 18 transcriptoproteome of liver tissue and HepG2 Cells and targeted proteome mapping in depleted plasma: Update 2013" [2], supporting the accompanying publication "State of the Chromosome 18-centric HPP in 2016: Transcriptome and Proteome Profiling of Liver Tissue and HepG2 Cells" [3], and are deposited at the ProteomeXchange via the PASSEL repository with the dataset identifier PASSEL: PASS00697 for liver and HepG2 cell line.
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http://dx.doi.org/10.1016/j.dib.2016.07.034DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4995477PMC
September 2016

Chemical modifications of amyloid-β(1-42) have a significant impact on the repertoire of brain amyloid-β(1-42) binding proteins.

Biochimie 2016 Sep-Oct;128-129:55-8. Epub 2016 Jul 8.

Engelhardt Institute of Molecular Biology, Russian Academy of Sciences, Vavilov Street 32, Moscow 119991, Russia.

The amyloid-β peptide(1-42) (Aβ) is a key player in the development and progression of Alzheimer's disease (AD). Although much attention is paid to its role in formation of extracellular amyloid plaques and protein aggregates as well as to corresponding mechanisms of their toxicity, good evidence exists that intracellular Aβ can accumulate intraneuronally and interact with intracellular target proteins. However, intracellular Aβ binding proteins as well as conditions favoring their interactions with Aβ are poorly characterized. In this study we have investigated the effect of two known pathogenic Aβ modifications, isomerization of Asp7 and phosphorylation of Ser8, on the proteomic profiles of mouse brain Aβ binding proteins. Phosphorylation of Ser8 and especially isomerization of Asp7 significantly extended the repertoire of mouse brain Aβ binding proteins. However, there were 61 proteins, common for three types of the affinity ligands. They obviously represent potential targets for direct interaction with all Aβ species. Taking into consideration spontaneous mode of Asp7 isomerization and reports on initial accumulation of phosphorylated Aβ species inside neurons it is reasonable to suggest that these modifications of intracellular Aβ peptides causing the significant increase in the repertoire of Aβ binding proteins represent a primary pathogenic effect that precedes formation of extracellular pathogenic oligomerization/aggregation of Aβ peptides well described in the literature.
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http://dx.doi.org/10.1016/j.biochi.2016.07.001DOI Listing
January 2017

The Size of the Human Proteome: The Width and Depth.

Int J Anal Chem 2016 19;2016:7436849. Epub 2016 May 19.

Institute of Biomedical Chemistry, Moscow 119121, Russia.

This work discusses bioinformatics and experimental approaches to explore the human proteome, a constellation of proteins expressed in different tissues and organs. As the human proteome is not a static entity, it seems necessary to estimate the number of different protein species (proteoforms) and measure the number of copies of the same protein in a specific tissue. Here, meta-analysis of neXtProt knowledge base is proposed for theoretical prediction of the number of different proteoforms that arise from alternative splicing (AS), single amino acid polymorphisms (SAPs), and posttranslational modifications (PTMs). Three possible cases are considered: (1) PTMs and SAPs appear exclusively in the canonical sequences of proteins, but not in splice variants; (2) PTMs and SAPs can occur in both proteins encoded by canonical sequences and in splice variants; (3) all modification types (AS, SAP, and PTM) occur as independent events. Experimental validation of proteoforms is limited by the analytical sensitivity of proteomic technology. A bell-shaped distribution histogram was generated for proteins encoded by a single chromosome, with the estimation of copy numbers in plasma, liver, and HepG2 cell line. The proposed metabioinformatics approaches can be used for estimation of the number of different proteoforms for any group of protein-coding genes.
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http://dx.doi.org/10.1155/2016/7436849DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4889822PMC
June 2016

Exome-based proteogenomics of HEK-293 human cell line: Coding genomic variants identified at the level of shotgun proteome.

Proteomics 2016 07 21;16(14):1980-91. Epub 2016 Jun 21.

Institute of Biomedical Chemistry, Moscow, Russia.

Genomic and proteomic data were integrated into the proteogenomic workflow to identify coding genomic variants of Human Embryonic Kidney 293 (HEK-293) cell line at the proteome level. Shotgun proteome data published by Geiger et al. (2012), Chick et al. (2015), and obtained in this work for HEK-293 were searched against the customized genomic database generated using exome data published by Lin et al. (2014). Overall, 112 unique variants were identified at the proteome level out of ∼1200 coding variants annotated in the exome. Seven identified variants were shared between all the three considered proteomic datasets, and 27 variants were found in any two datasets. Some of the found variants belonged to widely known genomic polymorphisms originated from the germline, while the others were more likely resulting from somatic mutations. At least, eight of the proteins bearing amino acid variants were annotated as cancer-related ones, including p53 tumor suppressor. In all the considered shotgun datasets, the variant peptides were at the ratio of 1:2.5 less likely being identified than the wild-type ones compared with the corresponding theoretical peptides. This can be explained by the presence of the so-called "passenger" mutations in the genes, which were never expressed in HEK-293 cells. All MS data have been deposited in the ProteomeXchange with the dataset identifier PXD002613 (http://proteomecentral.proteomexchange.org/dataset/PXD002613).
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http://dx.doi.org/10.1002/pmic.201500349DOI Listing
July 2016
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