scholarly journals 2D-ELECTROPHORESIS IN PROTEIN SPECTRUM CONSTRUCTION FOR EXOPROTEIN FRACTIONS OF PLAGUE AND CHOLERA AGENTS

2017 ◽  
pp. 40-44
Author(s):  
T. A. Polunina ◽  
S. P. Zadnova ◽  
Ya. M. Krasnov
Keyword(s):  
2D Electrophoresis ◽  
Protein Spectrum

Cross-Linked αsγt-Chain Hybrids in Plasma Clots Studied by 1D- and 2D Electrophoresis and Western Blotting

1993 ◽  
Vol 70 (03) ◽  
pp. 438-442 ◽  
Author(s):  
B Grøn ◽  
C Filion-Myklebust ◽  
S Bjørnsen ◽  
P Haidaris ◽  
F Brosstad
Keyword(s):  
Molecular Weight ◽  
Monoclonal Antibodies ◽  
Human Plasma ◽  
Western Blotting ◽  
Isoelectric Point ◽  
Factor Xiii ◽  
Cross Linking ◽  
2D Electrophoresis ◽  
Complex Phenomenon ◽  
Fibrinopeptide A

SummaryFibrinogen and fibrin related chains in reduced human plasma as well as the bonds interlinking partially cross-linked fibrin from plasma clots have been studied by means of 1D- and 2D electrophoresis and Western blotting. Immunovisualization of reduced plasma or partially cross-linked fibrin with monoclonal antibodies specific for the α-chains or the γ-chains have shown that several bands represent material belonging to both chains. In order to decide whether these bands constitute αγ-chain hybrids or superimposed α- and γ-chain dimers, the cross-linked material was separated according to both isoelectric point (pI) and molecular weight (MW) using Pharmacia’s Multiphor II system. Western blotting of the second dimension gels revealed that partially cross-linked fibrin contains αsγt-chain hybrids and γ- polymers, in addition to the well-known γ-dimers and α-polymers. The main αsγt-chain hybrid has a pI between that of the α- and the γ-chains, a MW of about 200 kDa and contains Aα-chains with intact fibrinopeptide A (FPA). It was also observed that soluble fibrinogen/fibrin complexes as well as partially cross-linked fibrin contain degraded α-dimers with MWs close to the γ-dimers. These findings demonstrate that factor XIII-catalyzed cross-linking of fibrin is a more complex phenomenon than earlier recognized.

scholarly journals Proteomic analysis identifies deregulated metabolic and oxidative-associated proteins in Italian intrahepatic cholangiocarcinoma patients

BMC Cancer ◽  
2021 ◽  
Vol 21 (1) ◽  
Author(s):  
Giuliana Cavalloni ◽  
Caterina Peraldo-Neia ◽  
Annamaria Massa ◽  
Carlo Bergamini ◽  
Alessandro Trentini ◽  
...  
Keyword(s):  
Intrahepatic Cholangiocarcinoma ◽  
Cell Structure ◽  
Duct Cell ◽  
Molecular Classification ◽  
Antioxidant Systems ◽  
2D Electrophoresis ◽  
Proteomic Data ◽  
Formalin Fixed Paraffin ◽  
Formalin Fixed Paraffin Embedded ◽  
Associated Proteins

Abstract Background Cholangiocarcinoma (CCA) is an aggressive disease with poor prognosis. A molecular classification based on mutational, methylation and transcriptomic features could allow identifying tailored therapies to improve CCA patient outcome. Proteomic remains partially unexplored; here, we analyzed the proteomic profile of five intrahepatic cholangiocarcinoma (ICC) derived from Italian patients undergone surgery and one normal bile duct cell line. Methods Proteome profile was investigated by using 2D electrophoresis followed by Mass Spectrometry (MS). To validate proteomic data, the expression of four overexpressed proteins (CAT, SOD, PRDX6, DBI/ACBP) was evaluated by immunohistochemistry in an independent cohort of formalin fixed, paraffin-embedded (FFPE) ICC tissues. We also compared proteomic data with those obtained by transcriptomic profile evaluated by microarray analysis of the same tissues. Results We identified 19 differentially expressed protein spots, which were further characterized by MS; 13 of them were up- and 6 were down-regulated in ICC. These proteins are mainly involved in redox processes (CAT, SODM, PRDX2, PRDX6), in metabolism (ACBP, ACY1, UCRI, FTCD, HCMS2), and cell structure and organization (TUB2, ACTB). CAT is overexpressed in 86% of patients, PRDX6 in 73%, SODM in 100%, and DBI/ACBP in 81% compared to normal adjacent tissues. A concordance of 50% between proteomic and transcriptomic data was observed. Conclusions This study pointed out that the impairment of the metabolic and antioxidant systems, with a subsequent accumulation of free radicals, might be a key step in CCA development and progression.

scholarly journals Proteome Analysis of Date Palm (Phoenix dactylifera L.) under Severe Drought and Salt Stress

2016 ◽  
Vol 2016 ◽  
pp. 1-8 ◽  
Author(s):  
Haddad A. El Rabey ◽  
Abdulrahman L. Al-Malki ◽  
Khalid O. Abulnaja
Keyword(s):  
Salt Stress ◽  
Drought Stress ◽  
Drought Tolerance ◽  
Date Palm ◽  
Proteome Analysis ◽  
Rubisco Activase ◽  
2D Electrophoresis ◽  
Severe Drought ◽  
Protein Abundance ◽  
Low Protein

Date palm cultivars differently tolerate salinity and drought stress. This study was carried out to study the response of date palm to severe salinity and drought based on leaf proteome analysis. Eighteen-month-old date palm plants were subjected to severe salt (48 g/L NaCl) and drought (82.5 g/L PEG or no irrigation) conditions for one month. Using a protein 2D electrophoresis method, 55 protein spots were analyzed using mass spectrometry. ATP synthase CF1 alpha chains were significantly upregulated under all three stress conditions. Changes in the abundance of RubisCO activase and one of the RubisCO fragments were significant in the same spots only for salt stress and drought stress with no irrigation, and oxygen-evolving enhancer protein 2 was changed in different spots. Transketolase was significantly changed only in drought stress with PEG. The expression of salt and drought stress genes of the chosen protein spots was either overexpressed or downexpressed as revealed by the high or low protein abundance, respectively. In addition, all drought tolerance genes due to no irrigation were downregulated. In conclusion, the proteome analysis of date palm under salinity and drought conditions indicated that both salinity and drought tolerance genes were differentially expressed resulting in high or low protein abundance of the chosen protein spots as a result of exposure to drought and salinity stress condition.

Fluoride exposure inhibits protein expression and enzyme activity in the lung-stage larvae ofAscaris suum

Parasitology ◽  
2006 ◽  
Vol 133 (4) ◽  
pp. 497-508 ◽  
Author(s):  
M. K. ISLAM ◽  
T. MIYOSHI ◽  
M. YAMADA ◽  
M. A. ALIM ◽  
X. HUANG ◽  
...  
Keyword(s):  
Protein Expression ◽  
Expression Profiles ◽  
Immunoblot Analysis ◽  
Specific Protein ◽  
Inorganic Pyrophosphatase ◽  
2D Electrophoresis ◽  
Peptide Mass ◽  
Enzyme Families ◽  
Protein Expression Profiles ◽  
Peptide Mass Spectrometry

Sodium fluoride (NaF) is an anion that has been previously shown to block the moulting process ofAscaris suumlarvae. This study describes moulting and development-specific protein expression profiles ofA. suumlung-stage L3 (AsLL3) following NaF exposure. AsLL3s cultured in the presence or absence of NaF were prepared for protein analysis using two-dimensional (2D) electrophoresis. NaF exposure inhibited at least 22 proteins in AsLL3 compared with moulted larvae (i.e. AsLL4). A further comparison of AsLL4 with those of pre-cultured AsLL3 and NaF-exposed AsLL3 revealed 8 stage-specifically and 4 over-expressed proteins. Immunoblot analysis revealed an inhibition by NaF of 19 immunoreactive proteins. Enzyme assay and immunochemical data showed an inhibition of the moulting-specific inorganic pyrophosphatase activity by 41% and a decreased expression in NaF-treated larvae, indicating its significance in the moulting process. A protein spot associated with NaF inhibition was isolated and identified by peptide mass spectrometry and bioinformatics approaches to be a member of 3–hydroxyacyl–CoA dehydrogenase/short-chain dehydrogenase enzyme families. These results have implications for the identification of proteins specific to the moulting process as potential chemotherapeutic targets.

On indications for the use of diphenhydramine and pipolfen for the treatment of patients with peptic ulcer disease

1980 ◽  
Vol 61 (5) ◽  
pp. 55-56
Author(s):  
D. G. Ulmetieva
Keyword(s):  
Peptic Ulcer ◽  
Peptic Ulcer Disease ◽  
Cholinergic Activity ◽  
Passive Hemagglutination ◽  
Ulcer Disease ◽  
Protein Spectrum

The dynamics of the passive hemagglutination reaction (RPHA) of the P reaction of leukocytolysis in persons suffering from peptic ulcer disease in the stage of relapse and remission was investigated. In parallel, the study of cholinergic activity and protein spectrum of blood was carried out.

Abstract 238: Incubation of MDCO216 With Human Serum or Plasma Potentiates ABCA1-Mediated Cholesterol Efflux Capacity, Increases Prebeta-1 HDL and Causes a Shift From Small to Large Alpha HDL Particles

2014 ◽  
Vol 34 (suppl_1) ◽  
Author(s):  
Herman J Kempen ◽  
Dorota B Schranz ◽  
Bela Asztalos ◽  
Elias Jeyarajah ◽  
James Otvos ◽  
...  
Keyword(s):  
Human Serum ◽  
Human Plasma ◽  
Cholesterol Efflux ◽  
Time Dependent ◽  
Plaque Burden ◽  
Strong Increase ◽  
2D Electrophoresis ◽  
Cholesterol Efflux Capacity ◽  
J774 Cells ◽  
Synergistic Increase

MDCO216 is a complex of dimeric apoA-IMilano and POPC, shown to reduce atherosclerotic plaque burden. Here we studied the effect of incubation of human plasma or serum with MDCO216 on cholesterol efflux capacity from J774 cells, on prebeta-1 HDL and various HDL subfractions. At clinically relevant concentrations MDCO216 by itself markedly stimulates global and ABCA1-mediated cholesterol efflux. When incubated with human serum a time-dependent synergistic increase of the ABCA1 mediated efflux capacity is observed. Using the Sekisui ELISA for prebeta-1 HDL, MDCO216 itself is poorly detected. Prebeta-1 HDL is rapidly lost when human plasma alone is incubated at 37 o C. However, incubation of human plasma with MDCO216 at 37 o C causes a robust generation of new prebeta-1 HDL. 2D-Electrophoresis followed by immunoblotting with a general apoA-I antibody that also detects apoA-IM confirms the increase in prebeta-1 HDL (having a different mobility than pure MDCO216 particles), and shows a concomitant disappearance of alpha-3 HDL, alpha-4 HDL and MDCO216, and an increase in alpha-1 and alpha-2 HDL. NMR analysis of plasma incubated with MDCO216 at 47 o C confirms very rapid disappearance of small HDL and increase of medium and large HDL particles. In conclusion, incubation of human plasma or serum with MDCO216 strongly enhances ABCA1-mediated cholesterol efflux capacity, causes a strong increase of prebeta-1 HDL and drastically changes HDL subfraction distribution, consistent with anti-atherosclerotic activity.

Oocyte proteomics: localisation of mouse zona pellucida protein 3 to the plasma membrane of ovulated mouse eggs

2004 ◽  
Vol 16 (2) ◽  
pp. 69 ◽  
Author(s):  
S. A. Coonrod ◽  
M. E. Calvert ◽  
P. P. Reddi ◽  
E. N. Kasper ◽  
L. C. Digilio ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Indirect Immunofluorescence ◽  
Zona Pellucida ◽  
Surface Expression ◽  
Surface Proteins ◽  
2D Electrophoresis ◽  
Dependent Manner ◽  
Two Dimensional ◽  
Phase Partitioning ◽  
Proteomic Approach

In order to gain a deeper understanding of the molecular underpinnings of sperm–egg interaction and early development, we have used two-dimensional (2D) electrophoresis, avidin blotting and tandem mass spectrometry to identify, clone and characterise abundant molecules from the mouse egg proteome. Two-dimensional avidin blots of biotinylated zona-free eggs revealed an abundant approximately 75-kDa surface-labelled heterogeneous protein possessing a staining pattern similar to that of the zona pellucida glycoprotein, mouse ZP3 (mZP3). In light of this observation, we investigated whether mZP3 specifically localises to the plasma membrane of mature eggs. Zona pellucidae of immature mouse oocytes and mature eggs were removed using acid Tyrode’s solution, chymotrypsin or mechanical shearing. Indirect immunofluorescence using the mZP3 monoclonal antibody (mAb) IE-10 demonstrated strong continuous staining over the entire surface of immature oocytes and weak microvillar staining on ovulated eggs, regardless of the method of zona removal. Interestingly, in mature eggs, increased fluorescence intensity was observed following artificial activation and fertilisation, whereas little to no fluorescence was observed in degenerated eggs. The surface localisation of ZP3 on mature eggs was supported by the finding that the IE-10 mAb immunoprecipitated an approximate 75-kDa protein from lysates of biotinylated zona-free eggs. To further investigate the specificity of the localisation of mZP3 to the oolemma, indirect immunofluorescence was performed using the IE-10 mAb on both CV-1 and CHO cells transfected with full-length recombinant mZP3 (re-mZP3). Plasma membrane targeting of the expressed re-mZP3 protein was observed in both cell lines. The membrane association of re-mZP3 was confirmed by the finding that biotinylated re-mZP3 (approximately 75 kDa) is immunoprecipitated from the hydrophobic phase of Triton X-114 extracts of transfected cells following phase partitioning. Immunoprecipitation assays also demonstrated that surface re-mZP3 was released from transfected CV-1 in a time-dependent manner. These results demonstrate that ZP3 is specifically associated with the surface of mature eggs and its subsequent release from the cell surface may represent one mechanism by which ZP3 is secreted. Furthermore, the increase in ZP3 surface expression following fertilisation suggests that ZP3 may have a functional role during sperm–oolemma binding and fusion. These results also validate the usefulness of using the 2D proteomic approach to identify and characterise egg-surface proteins.

scholarly journals Proteomic analysis of typical and genetically altered strains of Vibrio cholerae serogroup O1, biovar El Tor

2021 ◽  
Vol 97 (6) ◽  
pp. 578-586
Author(s):  
N. A. Plekhanov ◽  
S. P. Zadnova ◽  
A. A. Kritsky ◽  
T. A. Polunina ◽  
N. V. Kotova ◽  
...  
Keyword(s):  
Protein Expression ◽  
Vibrio Cholerae ◽  
Proteomic Analysis ◽  
Outer Membrane Proteins ◽  
Great Majority ◽  
Stress Factors ◽  
2D Electrophoresis ◽  
Oxidative Stresses ◽  
New Biomarkers ◽  
El Tor

Objective — comparative study of protein expression in typical and genetically altered Vibrio cholerae strains of O1 serogroup, biovar El Tor by means of proteomic analysis.Materials and methods. Clinical V. cholerae strains — typical strain, M106 (Astrakhan, 1970) and genetically altered one, M1509 (Moscow, 2012) — were used as model ones. Strains were cultivated in LB broth (pH7.2). Then, cell and exoprotein lysate fractions were obtained and investigated in 2D electrophoresis. Different protein stains were examined using mass spectrometry. Survivability of V. cholerae strains under osmotic and oxidative stresses was studied during incubation of the strains in 3 M NaCl solution or 20 mM H2O2 solution.Results and discussion. When analyzing cell lysates, significant differences in protein expression with known function between studied strains were not detected. The great majority of identified proteins in the lysates is functionally associated with carbohydrate metabolism, amino acid metabolism, and energy processes in a cell. At the same time, exoprotein fraction of M1509 genovariant contained increased amount of proteins (peroxidase, superoxide dismutase, thioredoxin, outer membrane proteins OmpW, OmpT) protecting the cells of cholera vibrio from effect of stress factors of the environment. Further study of the resistance to osmotic and oxidative stresses revealed better survivability in the genovariant when exposed to the stated factors.Conclusion. The data of proteomic analysis of the typical and genetically altered V. cholera strains, biovar El Tor, testify to high levels of expression of the proteins that provide for vibrio resistance to the effect of environmental stress factors in genovariants, which is possibly one of the causes of their wide dissemination. In addition, the results obtained will allow for identification of new biomarkers which can be used for differentiation of typical strains and genovariants of V. cholerae, biovar El Tor.

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