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2.
Zhonghua Yan Ke Za Zhi ; 55(10): 769-776, 2019 Oct 11.
Artigo em Chinês | MEDLINE | ID: mdl-31607066

RESUMO

Objective: To identify differentially expressed proteins between the patients with proliferative diabetic retinopathy (PDR) and vitreous floaters, and explore treatment target for PDR based on isobaric tags for relative and absolute quantification (iTRAQ) LC-MS/MS Proteomics. Method: Vitreous samples were collected from 28 eyes of patients with PDR and 4 eyes with vitreous floaters, which served as controls. For quantitative proteomics, vitreous samples were combined and proteins extracted and labeled with iTRAQ peptide-tagging reagents. Samples were fractionated by liquid chromatography (LC), analyzed by tandem mass spectrometry (MS/MS) and Gene Ontology (GO) analyses performed on differentially expressed proteins identified in the PDR samples. Results: In the PDR vitreous, 26 proteins were identified that were differentially expressed when compared to the controls. Of these, 7 showed a significant increase (P<0.05) and 19 a significant decrease (P<0.05)in expression in PDR patients. These included some high abundance proteins including Retinoic acid receptor reactive protein 2 (PDR 1=85.0, PDR 2=83.0, Control 1=119.6, Control 2=120.2, FC=0.710, P=0.001), Semaphorin-4B(PDR 1=64.4, PDR 2=68.8, Control 1=135.4, Control 2=146.0, FC=0.473, P=0.023), Apolipoprotein B (PDR 1=104.4, PDR 2=106.6, Control 1=89.0, Control 2=85.3, FC=1.211, P=0.024), and Heat shock protein 70 (PDR 1=69.3, PDR 2=75.0, Control 1=137.7, Control 2=138.3, FC=0.523, P=0.026), which are closely related to the pathological mechanism of PDR. GO analysis clustered the differentially expressed genes into three major functional domains: Biological Processes, Molecular Function and Cellular Component. Differential gene expression was found in the categories of cellular metabolism, organonitrogen compound and carbohydrate derivative metabolic processes, transferase activity and transmembrane signaling receptor activity. KEGG Pathway analysis indicate that Chemerin signaling through Akt, Sema4B signaling via PI3K, and HIF-1α signal pathways were all altered in the PDR samples. Conclusions: In this study we identified variations in expression of genes extensively involved in key biological processes in the retina including neovascularization, cellular metabolism and transmembrane signaling, which provide new insights into the pathophysiology of PDR. Extracellular matrix was degraded and endothelial cell migration was induced by Chemerin, in addition, the destruction of blood-retinal barrier and neuronal apoptosis were induced by ApoB. Chemerin and ApoB accelerated the development of PDR. Sema 4B participated in vascular protection, HSP70 conducted anti-apoptosis. These two cytokines protected the retinal neurovascular in PDR patients. Therefore, Chemerin, Sema 4B, ApoB and HSP70 may be the treatment target for PDR. (Chin J Ophthalmol, 2019, 55:769-776).


Assuntos
Diabetes Mellitus Tipo 2/complicações , Retinopatia Diabética/metabolismo , Peptídeos e Proteínas de Sinalização Intercelular/metabolismo , Proteoma/metabolismo , Proteômica/métodos , Corpo Vítreo/metabolismo , Biomarcadores/metabolismo , Estudos de Casos e Controles , Quimiocinas/metabolismo , Cromatografia Líquida/métodos , Diabetes Mellitus Tipo 2/metabolismo , Ensaio de Imunoadsorção Enzimática , Humanos , Proteoma/análise , Espectrometria de Massas em Tandem/métodos , Corpo Vítreo/química
3.
Se Pu ; 37(8): 853-862, 2019 Aug 08.
Artigo em Chinês | MEDLINE | ID: mdl-31642256

RESUMO

Diabetes is a systemic metabolic disorder syndrome, mainly characterized by hyperglycemia, and is associated with the dysfunction of various organs, such as liver, pancreas, intestine, adipose muscle tissue, kidney and brain. It has become a global epidemic disease that seriously threatens human health. Therefore, mapping the global molecular signatures of diabetes-related disease spectrum can provide more comprehensive data to understand early clinical diagnosis, molecular typing, and pathological processes involved in diabetes mellitus. In this study, we performed a quantitative differential analysis on the endogenous peptidome of the serum samples obtained from healthy, prediabetes and type 2 diabetes groups to explore the peptidomics evolution in the development of diabetes. Partial least squares-discriminant analysis (PLS-DA) was used for pattern recognition. A nonparametric test was examined to find out the significantly changed endogenous peptides. As a result, 690 serum endogenous peptides were identified totally, among which 163 endogenous peptides were statistically different among the three groups. This could be promising quantitative peptidomics data for early screening, diagnosis and molecular typing of type 2 diabetes mellitus.


Assuntos
Diabetes Mellitus Tipo 2/sangue , Peptídeos/sangue , Proteoma/análise , Humanos
4.
Nature ; 574(7776): 103-107, 2019 10.
Artigo em Inglês | MEDLINE | ID: mdl-31511700

RESUMO

The sequencing of ancient DNA has enabled the reconstruction of speciation, migration and admixture events for extinct taxa1. However, the irreversible post-mortem degradation2 of ancient DNA has so far limited its recovery-outside permafrost areas-to specimens that are not older than approximately 0.5 million years (Myr)3. By contrast, tandem mass spectrometry has enabled the sequencing of approximately 1.5-Myr-old collagen type I4, and suggested the presence of protein residues in fossils of the Cretaceous period5-although with limited phylogenetic use6. In the absence of molecular evidence, the speciation of several extinct species of the Early and Middle Pleistocene epoch remains contentious. Here we address the phylogenetic relationships of the Eurasian Rhinocerotidae of the Pleistocene epoch7-9, using the proteome of dental enamel from a Stephanorhinus tooth that is approximately 1.77-Myr old, recovered from the archaeological site of Dmanisi (South Caucasus, Georgia)10. Molecular phylogenetic analyses place this Stephanorhinus as a sister group to the clade formed by the woolly rhinoceros (Coelodonta antiquitatis) and Merck's rhinoceros (Stephanorhinus kirchbergensis). We show that Coelodonta evolved from an early Stephanorhinus lineage, and that this latter genus includes at least two distinct evolutionary lines. The genus Stephanorhinus is therefore currently paraphyletic, and its systematic revision is needed. We demonstrate that sequencing the proteome of Early Pleistocene dental enamel overcomes the limitations of phylogenetic inference based on ancient collagen or DNA. Our approach also provides additional information about the sex and taxonomic assignment of other specimens from Dmanisi. Our findings reveal that proteomic investigation of ancient dental enamel-which is the hardest tissue in vertebrates11, and is highly abundant in the fossil record-can push the reconstruction of molecular evolution further back into the Early Pleistocene epoch, beyond the currently known limits of ancient DNA preservation.


Assuntos
DNA Antigo/análise , Esmalte Dentário/metabolismo , Fósseis , Perissodáctilos/classificação , Perissodáctilos/genética , Filogenia , Proteoma/genética , Proteômica , Motivos de Aminoácidos , Sequência de Aminoácidos , Animais , Teorema de Bayes , História Antiga , Humanos , Masculino , Perissodáctilos/metabolismo , Fosforilação/genética , Proteoma/análise
5.
Mol Biol (Mosk) ; 53(4): 685-691, 2019.
Artigo em Russo | MEDLINE | ID: mdl-31397442

RESUMO

In the past decade, mass spectrometry studies of skeletal muscles have become common. In this tissue, the abundance of several contractile proteins significantly limits the depth of the panoramic proteome analysis. The use of isobaric labels allows improving assessment of the changes in the protein content, while analyzing up to 10 samples in a single run. Here we present the results of a comparative study of various methods for the fractionation of skeletal muscle peptides labeled with an isobaric label iTRAQ. Samples from m. vastus lateralis of eight young males were collected with a needle biopsy. After digestion into peptides and labeling, the preparations were carried out according to three different protocols: (1) peptide purification, HPLC-MS/MS; (2) peptide purification, isoelectric focusing, HPLC-MS/MS; (3) high pH reverse-phase LC fractionation, HPLC-MS/MS. Fractionation of labeled peptides by high pH reverse-phase LC was the optimal strategy for increasing the depth of the proteome analysis. This approach, in addition to contractile and mitochondrial proteins, allowed us to detect a variety of regulatory molecules, including the nucleic acids binding the proteins, chaperones, receptors, and transcription factors.


Assuntos
Métodos Analíticos de Preparação de Amostras/métodos , Músculo Esquelético/química , Músculo Esquelético/metabolismo , Proteoma/análise , Proteômica/métodos , Coloração e Rotulagem/métodos , Humanos , Focalização Isoelétrica , Proteoma/química , Espectrometria de Massas em Tandem
6.
Biomed Khim ; 65(4): 294-305, 2019 Jun.
Artigo em Russo | MEDLINE | ID: mdl-31436170

RESUMO

HL-60 promyelocytic cells are a widely used as a model for studying induced granulocytic differentiation. Investigation of proteins of the nuclear fraction, particularly transcription factors, is necessary for a better understanding of molecular mechanisms of cell maturation. Mass spectrometry is a powerful tool for analyzing a proteome due to its high sensitivity, specificity and performance. In this paper, using the selected reaction monitoring (SRM) method, we have assessed the levels of RBPJ, STAT1, CEBPB, CASP3, VAV1, PRKDC, PARP1 and UBC9 nuclear proteins isolated using hypertonic buffer, detergents (sodium dodecyl sulfate (SDS), sodium deoxycholate (DOC) and fissionable detergent ProteaseMAX™) and using centrifugation in a sucrose density gradient. The minimum and maximum protein content was 1.13±0.28 and 14.34±1.63 fmol/mkg of total protein for the transcription factor RBPJ and ubiquitin-protein ligase type I UBC9, respectively. According to the results of shotgun mass spectrometric analysis of nuclear fractions, 2356 proteins were identified, of which 106 proteins were annotated as transcription factors. 37 transcription factors were uniquely identified in the fraction obtained by centrifugation in a sucrose density gradient, while only 9 and 8 transcription factors were uniquely identified in the nuclear fractions obtained using hypertonic buffer and detergents, respectively. The transcription factors identified in the HL-60 cell line represent regulatory molecules; their directed profiling under the influence of differentiation inducers, will shed light on the mechanism of granulocyte maturation.


Assuntos
Proteínas Nucleares/análise , Proteoma/análise , Proteômica , Fatores de Transcrição/análise , Células HL-60 , Humanos , Espectrometria de Massas
7.
Nat Methods ; 16(9): 809-812, 2019 09.
Artigo em Inglês | MEDLINE | ID: mdl-31406385
8.
Nat Methods ; 16(8): 703-706, 2019 08.
Artigo em Inglês | MEDLINE | ID: mdl-31363206

RESUMO

Proteins can be phosphorylated at neighboring sites resulting in different functional states, and studying the regulation of these sites has been challenging. Here we present Thesaurus, a search engine that detects and quantifies phosphopeptide positional isomers from parallel reaction monitoring and data-independent acquisition mass spectrometry experiments. We apply Thesaurus to analyze phosphorylation events in the PI3K/AKT signaling pathway and show neighboring sites with distinct regulation.


Assuntos
Fosfopeptídeos/análise , Fosfoproteínas/análise , Proteoma/análise , Ferramenta de Busca/métodos , Células HeLa , Humanos , Proteínas Substratos do Receptor de Insulina/metabolismo , Isomerismo , Células MCF-7 , Fosfatidilinositol 3-Quinases/metabolismo , Fosforilação , Proteínas Proto-Oncogênicas c-akt/metabolismo , Transdução de Sinais , Espectrometria de Massas em Tandem
9.
Nat Methods ; 16(9): 894-901, 2019 09.
Artigo em Inglês | MEDLINE | ID: mdl-31384043

RESUMO

Mass spectrometry enables global analysis of posttranslationally modified proteoforms from biological samples, yet we still lack methods to systematically predict, or even prioritize, which modification sites may perturb protein function. Here we describe a proteomic method, Hotspot Thermal Profiling, to detect the effects of site-specific protein phosphorylation on the thermal stability of thousands of native proteins in live cells. This massively parallel biophysical assay unveiled shifts in overall protein stability in response to site-specific phosphorylation sites, as well as trends related to protein function and structure. This method can detect intrinsic changes to protein structure as well as extrinsic changes to protein-protein and protein-metabolite interactions resulting from phosphorylation. Finally, we show that functional 'hotspot' protein modification sites can be discovered and prioritized for study in a high-throughput and unbiased fashion. This approach is applicable to diverse organisms, cell types and posttranslational modifications.


Assuntos
Ensaios de Triagem em Larga Escala/métodos , Fosfoproteínas/análise , Fosfoproteínas/química , Processamento de Proteína Pós-Traducional , Proteoma/análise , Temperatura Ambiente , Células HeLa , Humanos , Fosforilação , Domínios e Motivos de Interação entre Proteínas , Estabilidade Proteica
10.
Nat Methods ; 16(9): 902-910, 2019 09.
Artigo em Inglês | MEDLINE | ID: mdl-31384044

RESUMO

We report a liquid chromatography coupled to tandem mass spectrometry O-glycoproteomics strategy using data-independent acquisition (DIA) mode for direct analysis of O-glycoproteins. This approach enables characterization of glycopeptides and structures of O-glycans on a proteome-wide scale with quantification of stoichiometries (though it does not allow for direct unambiguous glycosite identification). The method relies on a spectral library of O-glycopeptides; the Glyco-DIA library contains sublibraries obtained from human cell lines and human serum, and it currently covers 2,076 O-glycoproteins (11,452 unique glycopeptide sequences) and the 5 most common core1 O-glycan structures. Applying the Glyco-DIA library to human serum without enrichment for glycopeptides enabled us to identify and quantify 269 distinct glycopeptide sequences bearing up to 5 different core1 O-glycans from 159 glycoproteins in a SingleShot analysis.


Assuntos
Biologia Computacional/métodos , Simulação por Computador , Glicopeptídeos/análise , Glicoproteínas/análise , Polissacarídeos/análise , Proteoma/análise , Proteômica/métodos , Glicosilação , Humanos
11.
J Insect Sci ; 19(4)2019 Jul 01.
Artigo em Inglês | MEDLINE | ID: mdl-31343690

RESUMO

Bombyx mori (Lepidoptera: Bombycidae) is an important economic insect and a classic Lepidopteran model system. Although immune-related genes have been identified at a genome-wide scale in the silkworm, proteins involved in immune defense of the silkworm have not been comprehensively characterized. In this study, two types of bacteria were injected into the silkworm larvae, Gram-negative Escherichia coli (Enterobacteriales: Enterobacteriaceae), or Gram-positive Staphylococcus aureus (Bacillales: Staphylococcaceae). After injection, proteomic analyses of hemolymph were performed by liquid chromatography-tandem mass spectrometry. In total, 514 proteins were identified in the uninduced control group, 540 were identified in the E. coli-induced group, and 537 were identified in the S. aureus-induced group. Based on Uniprot annotations, 32 immunological recognition proteins, 28 immunological signaling proteins, and 21 immunological effector proteins were identified. We found that 127 proteins showed significant upregulation, including 10 immunological recognition proteins, 4 immunological signaling proteins, 11 immunological effector proteins, and 102 other proteins. Using real-time quantitative polymerase chain reaction in the fat body, we verified that immunological recognition proteins, signaling proteins, and effector proteins also showed significant increases at the transcriptional level after infection with E. coli and S. aureus. Five newly identified proteins showed upregulation at both protein and transcription levels after infection, including 30K protein, yellow-d protein, chemosensory protein, and two uncharacterized proteins. This study identified many new immune-related proteins, deepening our understanding of the immune defense system in B. mori. The data have been deposited to the iProX with identifier IPX0001337000.


Assuntos
Bombyx/genética , Bombyx/imunologia , Escherichia coli/fisiologia , Proteínas de Insetos/imunologia , Proteoma/imunologia , Staphylococcus aureus/fisiologia , Animais , Bombyx/crescimento & desenvolvimento , Bombyx/microbiologia , Proteínas de Insetos/análise , Larva/crescimento & desenvolvimento , Larva/imunologia , Larva/microbiologia , Proteoma/análise , Proteômica
12.
Parasit Vectors ; 12(1): 339, 2019 Jul 10.
Artigo em Inglês | MEDLINE | ID: mdl-31292008

RESUMO

BACKGROUND: The primary cause of parasitic gastroenteritis in small ruminants in temperate regions is the brown stomach worm, Teladorsagia circumcincta. Host immunity to this parasite is slow to develop, consistent with the ability of T. circumcincta to suppress the host immune response. Previous studies have shown that infective fourth-stage T. circumcincta larvae produce excretory-secretory products that are able to modulate the host immune response. The objective of this study was to identify immune modulatory excretory-secretory proteins from populations of fourth-stage T. circumcincta larvae present in two different host-niches: those associated with the gastric glands (mucosal-dwelling larvae) and those either loosely associated with the mucosa or free-living in the lumen (lumen-dwelling larvae). RESULTS: In this study excretory-secretory proteins from mucosal-dwelling and lumen-dwelling T. circumcincta fourth stage larvae were analysed using comparative 2-dimensional gel electrophoresis. A total of 17 proteins were identified as differentially expressed, with 14 proteins unique to, or enriched in, the excretory-secretory proteins of mucosal-dwelling larvae. One of the identified proteins, unique to mucosal-dwelling larvae, was a putative peroxiredoxin (T. circumcincta peroxiredoxin 1, Tci-Prx1). Peroxiredoxin orthologs from the trematode parasites Schistosoma mansoni and Fasciola hepatica have previously been shown to alternatively activate macrophages and play a key role in promoting parasite induced Th2 type immunity. Here we demonstrate that Tci-Prx1 is expressed in all infective T. circumcincta life-stages and, when produced as a recombinant protein, has peroxidase activity, whereby hydrogen peroxide (H2O2) is reduced and detoxified. Furthermore, we use an in vitro macrophage stimulation assay to demonstrate that, unlike peroxiredoxins from trematode parasites Schistosoma mansoni and Fasciola hepatica, Tci-Prx1 is unable to alternatively activate murine macrophage cells. CONCLUSIONS: In this study, we identified differences in the excretory-secretory proteome of mucosal-dwelling and lumen-dwelling infective fourth-stage T. circumcincta larvae, and demonstrated the utility of this comparative proteomic approach to identify excretory-secretory proteins of potential importance for parasite survival and/or host immune modulation.


Assuntos
Proteínas de Helminto/metabolismo , Peroxirredoxinas/metabolismo , Trichostrongyloidea/metabolismo , Animais , Eletroforese em Gel Bidimensional , Proteínas de Helminto/genética , Interações Hospedeiro-Parasita , Peróxido de Hidrogênio/metabolismo , Enteropatias Parasitárias , Larva/imunologia , Larva/metabolismo , Camundongos , Membrana Mucosa/parasitologia , Peroxirredoxinas/genética , Proteoma/análise , Proteômica , Ovinos/parasitologia , Doenças dos Ovinos/parasitologia , Trichostrongyloidea/imunologia
13.
Adv Exp Med Biol ; 1140: 541-561, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31347070

RESUMO

Proteomics involves large-scale comprehensive study of specific proteomes which have been widely used in the field of biomarker discovery, drug development, disease diagnosis and therapy. Comprehensive proteomics involve two or more proteomics approaches that are confirmatory, complementary, and/or synergistic. Obstructive sleep apnea (OSA) is a sleep disorder which causes respiratory cessation (due to upper airway collapse). Here we describe a comprehensive MS based label-free quantitative proteomic analysis of the OSA induced rat atria homogenates and matched controls by using 1 dimensional SDS PAGE (1-D PAGE) and 2 dimensional SDS PAGE (2-D PAGE) separation of the proteins, enzymatic digestion and analysis by nanoliquid chromatography tandem-mass spectrometry (LC-MS/MS). The outcomes from the 1D-PAGE and 2D-PAGE studies not only identified dysregulated proteins due to OSA, but also confirmed and complemented each other.


Assuntos
Átrios do Coração/metabolismo , Proteoma/análise , Proteômica , Apneia Obstrutiva do Sono , Animais , Cromatografia Líquida , Eletroforese em Gel de Poliacrilamida , Ratos , Espectrometria de Massas em Tandem
14.
Adv Exp Med Biol ; 1140: 585-600, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31347073

RESUMO

Major depressive disorder (MDD) is common. Despite numerous available treatments, many individuals fail to improve clinically. Diagnosis of MDD continues to be commonly accomplished via behavioral rather than biological methods. Biomarkers may provide objective diagnosis of MDD, and could include measurements of genes, proteins, and patterns of brain activity. Proteomic analysis and validation of biomarkers is less explored than other areas of biomarker research in MDD. Mass spectrometry (MS) is a comprehensive, unbiased means of proteomic analysis, which can be complemented by directed protein measurements, such as Western Blotting. Prior studies have focused on MS analysis of several human biomaterials in MDD, including human post-mortem brain, cerebrospinal fluid (CSF), blood components, and urine. Further studies utilizing MS and proteomic analysis in MDD may help solidify and establish biomarkers for use in diagnosis, identification of new treatment targets, and understanding of the disorder. A biomarker or a biomarker signature that facilitates a convenient and inexpensive predictive test for depression treatment response is highly desirable.


Assuntos
Biomarcadores/análise , Transtorno Depressivo Maior/diagnóstico , Espectrometria de Massas , Proteoma/análise , Humanos , Proteômica
15.
Nat Methods ; 16(8): 737-742, 2019 08.
Artigo em Inglês | MEDLINE | ID: mdl-31308550

RESUMO

Protein complexes are key macromolecular machines of the cell, but their description remains incomplete. We and others previously reported an experimental strategy for global characterization of native protein assemblies based on chromatographic fractionation of biological extracts coupled to precision mass spectrometry analysis (chromatographic fractionation-mass spectrometry, CF-MS), but the resulting data are challenging to process and interpret. Here, we describe EPIC (elution profile-based inference of complexes), a software toolkit for automated scoring of large-scale CF-MS data to define high-confidence multi-component macromolecules from diverse biological specimens. As a case study, we used EPIC to map the global interactome of Caenorhabditis elegans, defining 612 putative worm protein complexes linked to diverse biological processes. These included novel subunits and assemblies unique to nematodes that we validated using orthogonal methods. The open source EPIC software is freely available as a Jupyter notebook packaged in a Docker container (https://hub.docker.com/r/baderlab/bio-epic/).


Assuntos
Proteínas de Caenorhabditis elegans/metabolismo , Caenorhabditis elegans/metabolismo , Complexos Multiproteicos/isolamento & purificação , Complexos Multiproteicos/metabolismo , Mapeamento de Interação de Proteínas , Proteoma/análise , Software , Animais , Proteínas de Caenorhabditis elegans/isolamento & purificação
16.
Anal Chim Acta ; 1076: 82-90, 2019 Oct 17.
Artigo em Inglês | MEDLINE | ID: mdl-31203967

RESUMO

We present a method to preserve and process urine proteins for proteomic analysis in a filter aided sample preparation (FASP) format. The method combines concentration of urine proteins in ultrafiltration devices, their thermal stabilization, allowing long term storage of the samples, and filter aided sample preparation. Proteomes of four different urines were preserved during 48 h and 6 months using (i) the classic freeze preservation at -20 °C, (ii) snap-heated freeze-free preservation at laboratory temperature (20 °C) and (iii) snap-heated preservation at -60 °C. The three storage methods can effetely preserve the urine proteome for at least 6 months without significant alterations. Abundances of more than 500 proteins and specially 24 selected -cleared or -approved protein assayed in serum or plasma were found similar within the three preservation methods assessed. The new method here proposed dramatically simplifies the conditions for preserving the urine proteome for biobanks in terms of space and storage, including lowering the risks of sample degradation caused by misfunction of the freezer. Furthermore, the shipping of large number of samples can be made without the need of freezing. The application of the FASP format to isolate and preserve the proteins facilitates long-term storage and processing of proteome of urine samples.


Assuntos
Proteoma/química , Manejo de Espécimes/métodos , Urina/química , Lesão Renal Aguda/urina , Adulto , Biomarcadores/análise , Análise por Conglomerados , Calefação , Humanos , Biópsia Líquida , Masculino , Estudo de Prova de Conceito , Proteoma/análise , Proteoma/isolamento & purificação , Adulto Jovem
17.
Mol Biol (Mosk) ; 53(3): 524-528, 2019.
Artigo em Russo | MEDLINE | ID: mdl-31184618

RESUMO

Proteins with homo-repeats of more than 4 amino acid residues in length were examined to understand whether some splicing sites in pre-mRNA may be attributed to homo-repeats in human proteins. The human proteome was found to contain a total of 404 proteins with homo-repeats that account for at least one splicing site in pre-mRNA. Pre-mRNA splicing sites were more often found in the C-terminal part (67%) than in the middle orN-terminal part of a homo-repeat. Ten homo-repeats were identified to have two splicing sites per repeat. The repeats were lysine homo-repeats in all but one case.


Assuntos
Proteínas/análise , Proteínas/química , Precursores de RNA/genética , Sítios de Splice de RNA/genética , Sequências Repetitivas de Aminoácidos/genética , Humanos , Lisina/genética , Lisina/metabolismo , Proteínas/genética , Proteoma/análise , Proteoma/química , Proteoma/genética , Processamento de RNA/genética
18.
Food Chem ; 297: 125016, 2019 Nov 01.
Artigo em Inglês | MEDLINE | ID: mdl-31253287

RESUMO

To gain a better understanding concerning factors underlying carotenoid metabolism in banana pulp we investigated the carotenoid profile, metabolome, proteome and relative expression levels of carotegenesis-associated genes of fruit pulp in the two banana varieties ON and GN, with ON being characterized of high carotenoid accumulation. Results showed that high carotenoid content in banana pulp was well correlated with the relative expression of carotenogenesis-associated genes and the abundance of the corresponding proteins. An elevated accumulation of sugar metabolism-related compounds and a decreased amino acid accumulation were also observed in ON. Additionally proteins involved in the glycolytic pathway were more highly abundant in ON suggesting that this supports the higher accumulation of carotenoid in this genotype. We suggest that up-regulated expression of carotenogenesis-associated genes alongside elevated carbohydrate accumulation contribute to high carotenoid content in banana pulp, implying that a multi-target approach is necessary in order to improve carotenoid content in banana.


Assuntos
Carotenoides/análise , Metaboloma , Metabolômica/métodos , Musa/metabolismo , Proteômica/métodos , Aminoácidos/análise , Aminoácidos/metabolismo , Metabolismo dos Carboidratos , Carotenoides/metabolismo , Cromatografia Líquida de Alta Pressão , Análise Discriminante , Frutas/metabolismo , Cromatografia Gasosa-Espectrometria de Massas , Glicólise , Proteínas de Plantas/genética , Proteoma/análise , Espectrometria de Massas em Tandem
19.
Pol J Microbiol ; 68(2): 255-261, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31250596

RESUMO

The proteomes of outer membrane vesicles (OMVs) secreted by C. jejuni 81-176 strain, which was exposed to oxygen or antibiotic stress (polymyxin B), were characterized. We also assessed the OMVs production and their content in two mutated strains - ∆dsbI and ∆htrA. OMVs production was significantly increased under the polymyxin B stress and remained unaltered in all other variants. Interestingly, the qualitative load of OMVs was constant regardless of the stress conditions or genetic background. However, certain proteins exhibited notable quantitative changes, ranging from 4-fold decrease to 10-fold increase. Up- or downregulated proteins (e.g. major outer membrane protein porA, iron ABC transporter, serine protease- htrA, 60 kDa chaperonin-groL, enolase) represented various cell compartments (cytoplasm, periplasm, and membrane) and exhibited various functions; nevertheless, one common group was noted that consisted of components of flagellar apparatus, i.e., FlaA/B, FlgC/E, which were mostly upregulated. Some of these proteins are the putative substrates of DsbI protein. Further investigation of the regulation of C. jejuni OMVs composition and their role in virulence will allow a better understanding of the infectious process of C. jejuni.The proteomes of outer membrane vesicles (OMVs) secreted by C. jejuni 81­176 strain, which was exposed to oxygen or antibiotic stress (polymyxin B), were characterized. We also assessed the OMVs production and their content in two mutated strains ­ ∆dsbI and ∆htrA. OMVs production was significantly increased under the polymyxin B stress and remained unaltered in all other variants. Interestingly, the qualitative load of OMVs was constant regardless of the stress conditions or genetic background. However, certain proteins exhibited notable quantitative changes, ranging from 4-fold decrease to 10-fold increase. Up- or downregulated proteins (e.g. major outer membrane protein porA, iron ABC transporter, serine protease- htrA, 60 kDa chaperonin-groL, enolase) represented various cell compartments (cytoplasm, periplasm, and membrane) and exhibited various functions; nevertheless, one common group was noted that consisted of components of flagellar apparatus, i.e., FlaA/B, FlgC/E, which were mostly upregulated. Some of these proteins are the putative substrates of DsbI protein. Further investigation of the regulation of C. jejuni OMVs composition and their role in virulence will allow a better understanding of the infectious process of C. jejuni.


Assuntos
Proteínas de Bactérias/análise , Campylobacter jejuni/química , Campylobacter jejuni/efeitos dos fármacos , Vesículas Extracelulares/química , Deleção de Genes , Proteoma/análise , Estresse Fisiológico , Antibacterianos/farmacologia , Campylobacter jejuni/genética , Proteínas de Choque Térmico/deficiência , Oxirredutases/deficiência , Oxigênio/toxicidade
20.
Food Chem ; 295: 129-137, 2019 Oct 15.
Artigo em Inglês | MEDLINE | ID: mdl-31174741

RESUMO

Heat stress causes oxidative damage and quality reduction in poultry. Here, a tandem mass tag proteomic approach was applied to investigate the proteomic differences in duck meat from birds exposed to heat stress. Altogether 212 differential proteins were identified, including 178 down-regulated and 34 up-regulated proteins, compared to the control. Malondialdehyde and carbonyl content and cooking loss of the chest muscle significantly increased under heat stress. The proteomic analysis indicated that heat stress suppressed mitochondrial functions and respiratory chains, which might be responsible for the higher oxidation level. The results also revealed potential protective proteins involved in the defensive mechanisms against heat stress in duck muscles, such as sarcoplasmic/endoplasmic reticulum calcium ATPases, Mn-superoxide dismutase, heat shock protein family B member 7, methyltransferase like 21C, myosin-binding protein C, and carbonic anhydrase 3. These results provide potential targets for the research and identification of oxidative meat products due to heat stress.


Assuntos
Carne/análise , Estresse Oxidativo , Proteoma/análise , Proteômica/métodos , Animais , ATPases Transportadoras de Cálcio/genética , ATPases Transportadoras de Cálcio/metabolismo , Culinária , Patos/metabolismo , Proteínas de Choque Térmico HSP27/genética , Proteínas de Choque Térmico HSP27/metabolismo , Masculino , Músculo Esquelético/metabolismo , Mapas de Interação de Proteínas , Superóxido Dismutase/genética , Superóxido Dismutase/metabolismo
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