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The resistance of silkworms to Bombyx mori nuclear polyhedrosis virus (BmNPV) is controlled by a major dominant gene and multiple modifying genes. Given the presence of modified genes, it is difficult to determine the main gene by positional cloning. In this study, the main anti-BmNPV gene of BmNPV-resistant silkworm variety N was introduced into the susceptible variety Su to breed the near-isogenic line SuN with BmNPV resistance. The infection process of BmNPV in the hemolymph of Su and SuN was analyzed using the cell analysis system TissueFAXS PLUS. According to the law of infection and proliferation, hemolymph was extracted every 6 h for two-dimensional electrophoresis (2-DE) analysis and quantitative real-time polymerase chain reaction (qRT-PCR). Seven DEPs were found in comparisons between Su and SuN by 2-DE analysis. Among them, acid phosphatase, storage protein, and phenoloxidase can prevent pathogen invasion, which may play a role against BmNPV. Polyamine oxidase plays an important role in energy metabolism, which may be indirectly involved in the process of resisting BmNPV. Most of the transcriptional expression profiles of the seven DEPs were consistent with the 2-DE results. This study can provide a reference for the identification of anti-BmNPV genes and the breeding of BmNPV-resistant silkworm varieties. [ABSTRACT FROM AUTHOR]

This study aimed to assess the effects of low-temperature stress on two tomato cultivars (S-22 and PKM-1) treated with 24-epibrassinolide (EBL) and selenium (Se) by determining the changes in the proteomics profiles, growth biomarkers, biochemical parameters, and physiological functions. The growth parameters, photosynthetic traits, and activity of nitrate reductase in the S-22 and PKM-1 plants were markedly reduced by exposure to low temperatures. However, the combined application of EBL and Se under different modes significantly enhanced the aforementioned parameters under stress and non-stress conditions. Exposure to low temperatures increased the activities of the antioxidant enzymes (catalase, peroxidase, and superoxide dismutase) and the proline content of leaves, which were further enhanced by treatment with Se and EBL in both varieties. This research sheds light on the potential for employing exogenous EBL and Se as crucial biochemical tactics to assist tomato plants in surviving low-temperature stress. Moreover, the differentially expressed proteins that were involved in plant metabolism following the combined application of EBL and Se under low-temperature stress were additionally identified. Functional analysis revealed that the Q54YH4 protein plays an active role against plant stressors. The conserved regions in the protein sequences were analyzed for assessing the reliability of plant responses against the external application of EBL and Se under low temperatures. [ABSTRACT FROM AUTHOR]

BACKGROUND: Snail allergy is rare but can be fatal. Pila polita, a freshwater snail, was considered as a popular exotic food, particularly in tropical countries, and consumed in processed forms. Thus, the purpose of this study was to identify the major and cross‐reactive allergens of P. polita and to determine the impact of food processing on the allergen stability. RESULTS: Sodium dodecyl sulfate–polyacrylamide gel electrophoresis fractionated raw snail extract to approximately 24 protein bands, between 9 and 245 kDa. The prominent band at 33 kDa was detected in all raw and processed snail extracts. Immunoblotting tests of the raw extract demonstrated 19 immunoglobulin E (IgE)‐binding proteins, and four of them, at 30, 35, 42 and 49 kDa, were revealed as the major IgE‐binding proteins of P. polita. Matrix‐assisted laser desorption/ionization time‐of‐flight mass spectrometry identified the 49 and 42 kDa major allergens as actin, whereas the 30 and 35 kDa major allergens were identified as tropomyosin. Immunoblotting revealed that the raw snail had more allergenic proteins than the processed snail. The degree of allergenicity in decreasing order was raw > brine pickled> boiled > roasted > fried > vinegar pickled. The presence of cross‐reactivity between P. polita and the shellfish tested was exhibited with either no, complete, or partial inhibitions. CONCLUSION: Actin and tropomyosin were identified as the major and cross‐reactive allergens of P. polita among local patients with snail allergy. Those major allergens are highly stable to high temperatures, acidic pH, and high salt, which might played a crucial role in snail allergy in Malaysia. © 2023 Society of Chemical Industry. [ABSTRACT FROM AUTHOR]

Abstract Background Cystic echinococcosis (CE)/hydatidosis is an important zoonotic parasitic disease caused by the larval stage of Echinococcus granulosus. The disease is a major health problem all over the world. Finding specific and sensitive biomarkers for follow-up of CE in patients after surgery is essential. Using proteomics methods, the present study aimed to evaluate post-surgical treatment by finding probable biomarker/s in the serum of human lungs CE. Methods A total of 24 human sera were tested. These sera included eight confirmed lung/s CE patients sera before surgery (BS), eight sera 12 months post-surgery (12MPS) as well as eight control sera from healthy people. Proteomics methods including 2DE and LC–MS/MS were performed on the specimens followed by bioinformatics analysis. Differentially expressed proteins (DEP) were detected and, separately integrated with protein–protein interaction (PPI) data to construct the PPI network. Results A total of 171 protein spots were detected in three groups including BS, 12MPS, and control groups; of which a total of 106 DEP have been expressed based on fold changes > = 2 and p-value

Processing of secondary products after slaughter of farm animals is in demand. The pancreas is a rich source of bioactive protein substances, effective extraction of which is a serious problem today due to their aggregation. The aim of the work was to assess the extractivity of protein substances of the porcine pancreas using sodium chloride, trehalose, arginine, and combination of glycine and proline. The protein concentration was determined in the obtained extracts by the biuret reaction and their protein composition was assessed by densitometry of two-dimensional electropherograms using software ImageMaster™ 2D Platinum powered by Melanie 8.0. The results showed a positive effect of anti-aggregation agents on the release of protein substances into a solution. The highest protein concentration (33.36±0.64 g/l) was observed when adding 1М L-arginine; however, it was conditioned mainly by an increase in the content of three major protein fractions rather than by diversity of the protein composition. In general, the use of 0.9% NaCl as an extractive agent was quite effective, but selectivity to certain protein groups was observed for anti-aggregation agents such as sodium chloride, trehalose, arginine, glycine and proline, as well as their combination. The obtained results are important for intensifying extraction of protein substances including target ones with the subsequent application in different fields.

Abstract Learning to anticipate events based on the predictive relationship between an action and an outcome (operant conditioning) is a form of associative learning shared by humans and most of other living beings, including invertebrates. Several behavioral studies on the mechanisms of operant conditioning have included Melipona quadrifasciata, a honey bee that is easily manipulated due to lack of sting. In this work, brain proteomes of Melipona bees trained using operant conditioning and untrained (control) bees were compared by two-dimensional gel electrophoresis analysis within pI range of 3-10 and 4–7; in order to find proteins specifically related to this type of associative learning.One protein was detected with differential protein abundance in the brains of trained bees, when compared to not trained ones, through computational gel imaging and statistical analysis. This protein was identified by peptide mass fingerprinting and MS/MS peptide fragmentation using a MALDI-TOF/TOF mass spectrometer as one isoform of arginine kinase monomer, apparently dephosphorylated. Brain protein maps were obtained by 2-DE (Two-dimensional gel electrophoresis) from a total proteins and phosphoproteins extract of the bee Melipona quadrifasciata. One isoform of arginine kinase, probably a dephosphorylated isoform, was significantly more abundant in the brain of trained bees using operant conditioning. Arginine kinase has been reported as an important enzyme of the energy releasing process in the visual system of the bee, but it may carry out additional and unexpected functions in the bee brain for learning process.

Abstract This study aims to compare the grain protein profile of four Brazilian cowpea cultivars (BRS Aracê, BRS Itaim, BRS Pajeú, and BRS Xiquexique) by two-dimensional electrophoresis (2-DE) and principal component analysis (PCA). 2-DE efficiently separate cowpea protein profiles, showing high homogeneity among the four cultivars. In addition, the principal component analysis indicated that there is a difference in abundance of proteins among the cultivars. The cultivars BRS Aracê and BRS Xiquexique, both biofortified in iron and zinc, were separated from the cultivars BRS Itaim and BRS Pajeú. These results demonstrate that protein profiles can be used to discriminate cowpea varieties. Graphical Abstract

Proteome-based characterization of vegetative and flower bud formation was utilized to identify and differentiate protein species with significant variable abundance during floral transition in mango cv. Dashehari using 2DE and corroborating the identified protein spots using gene expression analysis. Total soluble proteins were phenol-extracted from the vegetative and floral flush of mango cv. Dashehari and separated on 2D gels at pH 4-7. The protein spots with variable intensity were identified through SameSpots software. The protein sequences of differentially accumulated spots were identified based on PI and MW using Citrus sinensis proteome isoelectric focusing database. Furthermore, these protein sequences were used to conduct (tBLASTn) against Mangifera indica to predict the protein. Real time gene expression was done to corroborate identified proteins. Total 301 spots were detected, out of which 16 were identified as differentially expressed based on P value (P≤0.05) and a 2-fold change. These 16 protein spots were identified on the basis of in silico comparative mapping protein against genome of mango and citrus, a close relative. They were classified into eight categories viz., transcriptional regulation, phenylpropanoid pathway and cell wall /cytoskeleton metabolism-related proteins, hormone signalling, flowering time, signal-transduction, transport and protein synthesis associated to flowering. Five genes coding for shortlisted proteins were used for validation of results using gene expression analysis. SAM (S adenosyl methionine synthase) was found to up-regulated in floral flush, involved in the biosynthesis of polyamines has an association with flowering and stress responses. Furthermore, ARF (Auxin Response Factor) and serine/threonine kinase gene members were also found to play a critical role in determining the floral development process in mango, consistent with results obtained through 2DE. Protein species putatively involved in phenylpropanoid pathway were also identified, showing the process of mango flowering from a new perspective beyond the conventional view. This study of flowering-related proteomics provides an overview of the biological pathways and regulatory mechanisms associated with the developmental physiology of flowering. [ABSTRACT FROM AUTHOR]

In this article, the proteomic profiles of pork by-products (snout, tongue, liver, kidney, spleen) were studied by comparative method on the first day and the fifth day of their storage. Two-dimensional electrophoresis according to O’Farrell was used for the aims of this article, while the results were further processed in ImageMaster software. Proteomic maps of by-products showed clear changes in protein composition after visualization and images analysis. There was a decrease and increase in manifestation intensity of some proteins. The study of the obtained electrophoregrams with the help of references resources allowed identifying various compounds in the by-products. 9 protein fractions with various intensity of manifestation were found on the day 1st and 5th. On the 1st day the following substances were intensively manifested: in the liver — glutathione peroxidase 4 (22.3 kDa), LEAP-2 (8.8 kDa); in the kidneys — quinone oxidoreductase (34.9 kDa); in the spleen — glycoprotein CD59 (13.7 kDa), in the patch — protein flint (49.07 kDa). It is noted that these proteins play their role in stopping certain processes in cells, like oxidation, microbial activity, and accumulation of toxic substances. These processes can worsen the quality of raw materials, and further lead to spoilage of the food product. On the 5th day of storage the highest intensity of manifestation of glyceraldehyde-3-phosphate dehydrogenase (35.8 kDa) in the liver was observed; superoxide dismutase [Cu-Zn] (15.8 kDa) was noted in the kidneys, colony-stimulating factor (16.2 kDa) was observed in the spleen and glutaredoxin –1 (11.8 kDa) in the tongue. In its turn, on the fifth day these chemical processes manifested themselves more intensely, as the fatty acids and glucose broke down. To obtain more accurate results, the proteins were compared by their volume. Among the identified fractions the highest expression was observed in LEAP 2 (8.8 kDa) on the first day, and in glyceraldehyde-3-phosphate dehydrogenase (35.8 kDa) on the fifth day. The least change in the intensity of manifestation was noted for superoxide dismutase [Cu-Zn] (15.8 kDa), which volume increased during storage by 13% for 5 days. The analysis of the obtained electrophoregrams allowed identifying various compounds, tracing the changes in the qualitative composition of protein in by-products during various periods of their storage. The obtained data demonstrate the transformation of protein molecules during storage, which makes it possible to determine the changes and quality of the food products.

The production of high-quality pork is closely related to the growth and development of muscle tissue. The present article provides a comparative proteomic research of l. dorsi, b. femoris, m. brachiocephalicus during the pigs’ growth and development (at age of 60 days and 180 days). This work was supported by data of electrophoretic methods: one-dimensional electrophoresis according to Laemmli with densitometric assessment in the ImageJ software and two-dimensional electrophoresis according to O’Farrell method with its further processing on the software ImageMaster. The mass spectrometric identification was conducted with the help of the high-performance liquid chromatography (HPLC) system connected to a mass spectrometer; further the data were interpreted by search algorithm Andromeda. When comparing frequency diagrams of one-dimensional electrophoregrams of all three muscle tissues of weaned pigs, the greatest difference was observed for the muscle sample l. dorsi. Comparison of diagrams of muscle tissue samples taken for mature pigs showed a great similarity of all three studied muscles samples. Within the framework of the research, the Fold indicator was calculated. The exceeding its value by more than 2 units is generally considered to be a statistically significant difference. When analyzing two-dimensional electrophoretograms of weaned pigs’ muscles, 18 protein fractions were revealed with Fold > 2. When examining the muscle tissue of mature pigs, 15 of those proteins were found; the differences were mostly detected in the minor protein fractions. The mass spectrometric analysis of the cut bands with well-pronounced differences from the onedimensional electrophoretogram revealed 214 proteins involved to a greater extent in cellular and metabolic processes, physical activity and localization. Growth and development protein — semaphorin‑6B (96.78 kDa) — was revealed in muscle tissue of l. dorsi, a. Also in l. dorsi and b. femoris the growth and development proteins were found: cadherin‑13 (78.23 kDa), cadherin‑7 (87.01 kDa), the F‑actin-cap protein beta subunit (30.66 kDa), and two uncharacterized proteins at 65.60 kDa and 63.88 kDa.

This study aimed to assess the effects of low-temperature stress on two tomato cultivars (S-22 and PKM-1) treated with 24-epibrassinolide (EBL) and selenium (Se) by determining the changes in the proteomics profiles, growth biomarkers, biochemical parameters, and physiological functions. The growth parameters, photosynthetic traits, and activity of nitrate reductase in the S-22 and PKM-1 plants were markedly reduced by exposure to low temperatures. However, the combined application of EBL and Se under different modes significantly enhanced the aforementioned parameters under stress and non-stress conditions. Exposure to low temperatures increased the activities of the antioxidant enzymes (catalase, peroxidase, and superoxide dismutase) and the proline content of leaves, which were further enhanced by treatment with Se and EBL in both varieties. This research sheds light on the potential for employing exogenous EBL and Se as crucial biochemical tactics to assist tomato plants in surviving low-temperature stress. Moreover, the differentially expressed proteins that were involved in plant metabolism following the combined application of EBL and Se under low-temperature stress were additionally identified. Functional analysis revealed that the Q54YH4 protein plays an active role against plant stressors. The conserved regions in the protein sequences were analyzed for assessing the reliability of plant responses against the external application of EBL and Se under low temperatures.

This study aims to compare the grain protein profile of four Brazilian cowpea cultivars (BRS Aracê, BRS Itaim, BRS Pajeú, and BRS Xiquexique) by two-dimensional electrophoresis (2-DE) and principal component analysis (PCA). 2-DE efficiently separate cowpea protein profiles, showing high homogeneity among the four cultivars. In addition, the principal component analysis indicated that there is a difference in abundance of proteins among the cultivars. The cultivars BRS Aracê and BRS Xiquexique, both biofortified in iron and zinc, were separated from the cultivars BRS Itaim and BRS Pajeú. These results demonstrate that protein profiles can be used to discriminate cowpea varieties. [ABSTRACT FROM AUTHOR]

Abstract Background Mangroves plants, an important wetland system in the intertidal shores, play a vital role in estuarine ecosystems. However, there is a lack of a very effective method for extracting protein from mangrove plants for proteomic analysis. Here, we evaluated the efficiency of three different protein extraction methods for proteomic analysis of total proteins obtained from mangrove plant Kandelia obovata leaves. Results The protein yield of the phenol-based (Phe-B) method (4.47 mg/g) was significantly higher than the yields of the traditional phenol (Phe) method (2.38 mg/g) and trichloroacetic acid-acetone (TCA-A) method (1.15 mg/g). The Phe-B method produced better two-dimensional electrophoresis (2-DE) protein patterns with high reproducibility regarding the number, abundance and coverage of protein spots. The 2-DE gels showed that 847, 650 and 213 unique protein spots were separated from the total K. obovata leaf proteins extracted by the Phe-B, Phe and TCA-A methods, respectively. Fourteen pairs of protein spots were randomly selected from 2-DE gels of Phe- and Phe-B- extracted proteins for identification by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF/TOF-MS) technique, and the results of three pairs were consistent. Further, oxygen evolving enhancer protein and elongation factor Tu could be observed in the 2-DE gels of Phe and Phe-B methods, but could only be detected in the results of the Phe-B methods, showing that Phe-B method might be the optimized choice for proteomic analysis. Conclusion Our data provides an improved Phe-B method for protein extraction of K. obovata and other mangrove plant tissues which is rich in polysaccharides and polyphenols. This study might be expected to be used for proteomic analysis in other recalcitrant plants.

Background and Aim: Carbapenem resistance has become a major obstacle in combating Acinetobacter baumannii infections. Although enzymatic degradation by β‐lactamases is the pivotal mechanism of carbapenem resistance, porin deficiency has also been implicated in the mechanism. In this study, outer membrane proteins (OMPs) pattern of a clinical multidrug‐resistant A. baumannii isolate were analysed in order to attain a deeper understanding of carbapenem‐resistance strategies. Methods: OMPs extracts, respectively, separated from carbapenem‐resistant and ‐susceptible clinical A. baumannii isolates were compared using two‐dimensional polyacrylamide gel electrophoresis. Differentially expressed proteins were identified by matrix‐assisted laser desorption/ionization time‐of‐flight (MALDI‐TOF). Results: Twenty‐three differently expressed proteins were identified between the resistant and susceptible isolates. Among them, six were annotated convincingly as OMPs in UniProt database. CarO was found absent from the resistant isolate and the expression levels of Omp33‐36 and Omp25 were significantly lower than that in the susceptible counterpart. Strikingly, a LysM domain/BON superfamily protein, which has been linked to carbapenem resistance in Klebsiella pneumoniae, was found underexpressed by tenfold in the resistant isolate. Conclusion: Our study verified some porins which have been proven to play an important role in bacterial resistance against carbapenems. Underexpression of the LysM domain/BON superfamily protein may indicate its possible engagement in bacterial drug resistance, but its actual role requires more investigation. [ABSTRACT FROM AUTHOR]

Iron (Fe) and manganese (Mn) are two essential elements for plants that compete for the same uptake transporters and show conflicting interactions at the regulatory level. In order to understand the differential response to both metal deficiencies in plants, two proteomic techniques (two-dimensional gel electrophoresis and label-free shotgun) were used to study the proteome profiles of roots from tomato plants grown under Fe or Mn deficiency. A total of 119 proteins changing in relative abundance were confidently quantified and identified, including 35 and 91 in the cases of Fe deficiency and Mn deficiency, respectively, with 7 of them changing in both deficiencies. The identified proteins were categorized according to function, and GO-enrichment analysis was performed. Data showed that both deficiencies provoked a common and intense cell wall remodelling. However, the response observed for Fe and Mn deficiencies differed greatly in relation to oxidative stress, coumarin production, protein, nitrogen, and energy metabolism. [ABSTRACT FROM AUTHOR]

The proteostasis system of animals, including various types of protein modification during the growth stage, leads to an almost incomprehensible number of possible forms of protein, and each can regulate numerous functions. In the presented work, the composition of muscle tissue protein from different portions of piglets was studied to understand the main muscle protein formation. Comparative analysis of weaned piglets' main muscle protein from l. dorsi, biceps femoris, and brachiocephalicus were analyzed using two-dimensional electrophoresis. Changes in the staining intensity of protein fractions inherent in different muscles were revealed. As part of this work, candidate groups of pig muscle proteins have been selected. Eleven protein spots were revealed for the longest muscle of the back, and seven for the biceps; the muscles of the neck are characterized by indicators of low protein fraction volume. Among the proteins found, myosin light chains, phosphoglycerate mutase, troponins, and adenylate kinase is most likely present. The obtained results of protein identification in muscle tissues, obtained during the intensive growth period, will allow a more detailed understanding of protein regulation, function, and interactions in complex biological systems, which will subsequently be significantly important for biomonitoring health and predicting farm animals productivity.

Natural rubber, an important industrial raw material with wide applications, is harvested in the form of latex (cytoplasm of rubber-producing laticifers) from Hevea brasiliensis (para rubber tree) by the way of tapping. Conspicuous stimulation on latex production is observed for the first few tappings conducted on virgin (untapped before) or resting (tapped before but no tapping for a period) rubber trees. To understand the underlying mechanisms, an integrative analysis of the latex transcriptome and proteome was conducted on virgin or resting Hevea trees for the first five tappings. A total of 505 non-redundant differentially expressed (DE) transcript-derived fragments (TDFs) were identified by silver-staining cDNA-AFLP, with 217 exhibiting patterns of upregulated, 180 downregulated and 108 irregularly-regulated. Meanwhile, 117 two dimensional gel electrophoresis DE-protein spots were isolated and subjected to mass spectrometry analysis, with 89 and 57 being successfully identified by MALDI-TOF and MALDI-TOF/TOF, respectively. About 72.5% DE-TDFs and 76.1% DE-proteins were functionally annotated and categorized. Noteworthily, most of the DE-TDFs implicated in sugar transport and metabolism as well as rubber biosynthesis were upregulated by the tapping treatment. The importance of sugar metabolism in harvesting-induced latex production was reinforced by the identification of abundant relevant DE-protein spots. About 83.8% of the randomly selected DE-TDFs were validated for expression patterns by semi-quantitative RT-PCR, and an 89.7% consistency for the 29 latex regeneration-related DE-TDFs examined by quantitative RT-PCR analysis. In brief, our results reveal extensive physiological and molecular changes in Hevea laticifers incurred by the tapping treatment, and the vast number of DE genes and proteins identified here contribute to unraveling the gene regulatory network of tapping-stimulated latex production. [ABSTRACT FROM AUTHOR]

Natural rubber, an important industrial raw material with wide applications, is harvested in the form of latex (cytoplasm of rubber-producing laticifers) from Hevea brasiliensis (para rubber tree) by the way of tapping. Conspicuous stimulation on latex production is observed for the first few tappings conducted on virgin (untapped before) or resting (tapped before but no tapping for a period) rubber trees. To understand the underlying mechanisms, an integrative analysis of the latex transcriptome and proteome was conducted on virgin or resting Hevea trees for the first five tappings. A total of 505 non-redundant differentially expressed (DE) transcript-derived fragments (TDFs) were identified by silver-staining cDNA-AFLP, with 217 exhibiting patterns of upregulated, 180 downregulated and 108 irregularly-regulated. Meanwhile, 117 two dimensional gel electrophoresis DE-protein spots were isolated and subjected to mass spectrometry analysis, with 89 and 57 being successfully identified by MALDI-TOF and MALDI-TOF/TOF, respectively. About 72.5% DE-TDFs and 76.1% DE-proteins were functionally annotated and categorized. Noteworthily, most of the DE-TDFs implicated in sugar transport and metabolism as well as rubber biosynthesis were upregulated by the tapping treatment. The importance of sugar metabolism in harvesting-induced latex production was reinforced by the identification of abundant relevant DE-protein spots. About 83.8% of the randomly selected DE-TDFs were validated for expression patterns by semi-quantitative RT-PCR, and an 89.7% consistency for the 29 latex regeneration-related DE-TDFs examined by quantitative RT-PCR analysis. In brief, our results reveal extensive physiological and molecular changes in Hevea laticifers incurred by the tapping treatment, and the vast number of DE genes and proteins identified here contribute to unraveling the gene regulatory network of tapping-stimulated latex production.

Background: Mangroves plants, an important wetland system in the intertidal shores, play a vital role in estuarine ecosystems. However, there is a lack of a very effective method for extracting protein from mangrove plants for proteomic analysis. Here, we evaluated the efficiency of three different protein extraction methods for proteomic analysis of total proteins obtained from mangrove plant Kandelia obovata leaves. Results: The protein yield of the phenol-based (Phe-B) method (4.47 mg/g) was significantly higher than the yields of the traditional phenol (Phe) method (2.38 mg/g) and trichloroacetic acid-acetone (TCA-A) method (1.15 mg/g). The Phe-B method produced better two-dimensional electrophoresis (2-DE) protein patterns with high reproducibility regarding the number, abundance and coverage of protein spots. The 2-DE gels showed that 847, 650 and 213 unique protein spots were separated from the total K. obovata leaf proteins extracted by the Phe-B, Phe and TCA-A methods, respectively. Fourteen pairs of protein spots were randomly selected from 2-DE gels of Phe- and Phe-B- extracted proteins for identification by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF/TOF-MS) technique, and the results of three pairs were consistent. Further, oxygen evolving enhancer protein and elongation factor Tu could be observed in the 2-DE gels of Phe and Phe-B methods, but could only be detected in the results of the Phe-B methods, showing that Phe-B method might be the optimized choice for proteomic analysis. Conclusion: Our data provides an improved Phe-B method for protein extraction of K. obovata and other mangrove plant tissues which is rich in polysaccharides and polyphenols. This study might be expected to be used for proteomic analysis in other recalcitrant plants. [ABSTRACT FROM AUTHOR]

Aims: Acinetobacter baumannii has been identified as one of the six most pathogenic bacteria that is the cause of most hospital bacterial infections according to Infectious Disease Society of America (IDSA). These nosocomial pathogens are notorious worldwide due to its ability in causing lethal infections among immunocompromised patients and its resistance to many strong antibiotics. This study aims to compare the expressed proteins of two A. baumannii strain, ATCC 19606 and a pathogenic clinically isolated strain known as AB-13. Methodology and results: AB-13 clinically strain was isolated from the lower respiratory tract of a patient with pneumonia. In this study, the proteomic profile of both ATCC 19606 and AB-13 are produced using 2-dimensional gel electrophoresis. The total protein contents were extracted, quantified and separated using 2-DE with a pH range of 4-7 to acquire the proteomic profile for comparison. The final analytical gel was analysed using Delta2D software and among the 324 protein spots successfully resolved, 10 spots exhibited signs of differential expression with 7 spots found to be downregulated and 3 spots upregulated (p< 0.01). These differences could signify the evolution AB-13 has undergone as it acquires traits ultimately aiding in its survivability, antimicrobial resistance and pathogenicity within varied environments especially during infections. Conclusion, significance and impact of study: These findings support the presence of variation in AB-13 from a proteomic perspective, highlighting the pathogen's evolution improving survivability and pathogenicity, warranting indepth exploration towards understanding A. baumannii virulence and pathogenicity. [ABSTRACT FROM AUTHOR]

Visceral leishmaniasis (VL), mainly caused by the Leishmania donovani parasitic infection, constitutes a potentially fatal disease, for which treatment is primarily dependent on chemotherapy. The emergence of a resistant parasite towards current antileishmanial agents and increasing reports of relapses are the major concerns. Detailed research on the molecular interaction at the host-parasite interface may provide the identification of the parasite and the host-related factors operating during disease development. Genomic and proteomic studies highlighted several essential secretory and cytosolic proteins that play vital roles during Leishmania pathogenesis. The aim of this study was to identify membrane proteins from the Leishmania donovani parasite and the host macrophage that interact with each other using 2-DE/MALDI-TOF/MS. We identified membrane proteins including activated protein C kinase, peroxidoxin, small myristoylated protein 1 (SMP-1), and cytochrome C oxidase from the parasite, while identifying filamin A interacting protein 1(FILIP1) and β-actin from macrophages. We further investigated parasite replication and persistence within macrophages following the macrophage-amastigote model in the presence or absence of withaferin (WA), an inhibitor of activated C kinase. WA significantly reduced Leishmania donovani replication within host macrophages. This study sheds light on the important interacting proteins for parasite proliferation and virulence, and the establishment of infection within host cells, which can be targeted further to develop a strategy for chemotherapeutic intervention [ABSTRACT FROM AUTHOR]

The diamondback moth (Plutella xylostella L.), is an economic pest of cruciferous plants worldwide, which causes great economic loss to cruciferous plants production. However, the pest has developed resistance to insecticides. One of such insecticides is chlorantraniliprole. The study of the mechanisms underlying resistance is key for the effective management of resistance. In this study, a comparative proteomics approach was used to isolate and identify various proteins that differed between chlorantraniliprole-susceptible and -resistant strains of P. xylostella. Eleven proteins were significantly different and were successfully identified by MALDI-TOF-MS. Metabolism-related proteins accounted for the highest proportion among the eleven different proteins. The function of the PxGST2L protein was validated by RNAi. Knockdown of PxGST2L reduced the GST activity and increased the toxicity of chlorantraniliprole to the diamondback moth. The resistance ratio of diamondback moth to chlorantraniliprole was reduced from 1029 to 505. The results indicated that PxGST2L is partly responsible for chlorantraniliprole insecticide resistance in DBM. Our finding contributes to the understanding of the mechanism underlying resistance to chlorantraniliprole in the DBM, to develop effective resistance management tactics. [ABSTRACT FROM AUTHOR]

Abstract Background The mesocotyl connects the coleoptilar node and the basal part of the seminal root of maize (Zea mays) seedling. The mesocotyl pushes the shoot of the seedling out of the soil during seed germination; thus, its growth is highly related to deep-sowing tolerance. Although many studies on the maize mesocotyl have been carried out at physiological and molecular levels, the proteomic changes associated with cellular and physiological activities during mesocotyl growth are still unknown. Results In the present study, the maize hybrid Zhengdan 958 was used to study mesocotyl growth and accompanying protein changes. The dark-grown etiolated mesocotyls exhibited a slow-fast-slow feature, with significant changes in the levels of indole-3-acetic acid (IAA) and cellulose and the activity of peroxidase (POD). In particular, POD activity increased with mesocotyl growth, showing higher activity at the mature (lower) end of the mesocotyl. For the proteomic analysis, soluble proteins were extracted from etiolated mesocotyls dark-grown for 48 h, 84 h, and 132 h, corresponding to the initial, rapid, and slow growth periods, respectively, and subjected to separation by two-dimensional gel electrophoresis (2-DE). As a result, 88 differentially abundant proteins (DAPs) were identified using MALDI-TOF-TOF analysis. At 48 h, most DAPs were stress proteins, heat shock proteins and storage proteins; at 84 h, oxidation/reduction proteins, carbohydrate biogenesis-related proteins and cytoskeleton-related proteins were highly accumulated; at 132 h, the most striking DAPs were those involved in the synthesis and modification of the cell wall and the biogenesis of carbohydrates. Gene ontology (GO) analysis showed that changes in the abundance and proportion of DAPs were consistent with cellular and physiological activities and biological processes during mesocotyl growth. The accumulation of nine DAPs of interest was verified by immunoblotting and RT-qPCR. Conclusions The present study revealed that the protein patterns in 2-D gels differed greatly with mesocotyl growth. At different growth periods, a specific set of DAPs participate in various biological processes and underlie the cellular and physiological activities of the mesocotyl. These results contributed to the understanding of mesocotyl growth and the cultivation of maize lines with deep-sowing tolerance.

Aspergillus fumigatus is one of the pathogenic filamentous fungi that could cause opportunistic infection, allergy and poisoning. Since, genetic sequencing of some of these fungi has been completed, identifying the protein profile of these fungus cells is necessary. One of these genes is the Annexin family. Annexin C4 is a new member of fungal annexins. This study investigates the effect of ANXC4 mutant gene in proteome profile of Aspergillus Fumigatus. Moreover, in order to enhance the power of protein complex separation, we used an optimized prefractionation method. Using reverse phase-high performance liquid chromatography eight fractions were separated. Then to confirm the protein concentration, each fraction was tested by SDS-PAGE. Protein profile of these fractions was analyzed using 2-DE, and Image Master software. Among the proteins identified statistically, two emerging proteins were observed. The results show that the expression of ANXC4 could affect the expression of some proteins in A.Fumigatus. To accurately identify these proteins, further experiments are needed including Mass spectrometry analysis.

BACKGROUND: Prepared chicken breast deterioration is a complex biochemical process, of which protein change is one of the main features. The present research focuses on the analysis of proteins related to the deterioration in quality of prepared chicken breast through differential proteomics analysis. RESULTS: The physicochemical indexes of prepared chicken breast showed that quality gradually decreased at the second week of refrigerated storage, while the deterioration of chicken breast meat was obvious at the third week. Three key time points of quality change were determined to be at 0th, 2th and 5th week, respectively. In addition, 39 differential proteins were successfully identified using two‐dimensional gel electrophoresis and matrix‐assisted laser desorption/ionization time‐of‐flight mass spectrometry. Most of the identified proteins showed significant differences in expression at the three key points of storage, of which actin, myosin, α‐1,4‐glucan phosphorylase, phosphoglucomutase 1, heat shock protein β‐1, tubulin β‐7 chain and skeletal muscle type tropomodulin (fragment) were closely related to the quality deterioration of prepared chicken breast, and thus potential indicator proteins to evaluate the quality of chicken breast. CONCLUSION: The current study indicated that the physicochemical quality of prepared breast notably changed during refrigerated storage. Three key time points of quality change in the storage process of prepared chicken breast were determined. Furthermore, differential proteomics identified the key proteins related to freshness, which provides a theoretical basis for exploring the mechanism of chicken breast deterioration. © 2020 Society of Chemical Industry [ABSTRACT FROM AUTHOR]

Candida auris is an emerging antifungal resistant human fungal pathogen increasingly reported in healthcare facilities. It persists in hospital environments, and on skin surfaces, and can form biofilms readily. Here, we investigated the cell surface proteins from C. auris biofilms grown in a synthetic sweat medium mimicking human skin conditions. Cell surface proteins from both biofilm and planktonic control cells were extracted with a buffer containing β-mercaptoethanol and resolved by 2-D gel electrophoresis. Some of the differentially expressed proteins were excised and identified by mass spectrometry. C. albicans orthologs Spe3p, Tdh3p, Sod2p, Ywp1p, and Mdh1p were overexpressed in biofilm cells when compared to the planktonic cells of C. auris. Interestingly, several proteins with zinc ion binding activity were detected. Nrg1p is a zinc-binding transcription factor that negatively regulates hyphal growth in C. albicans. C. auris does not produce true hypha under standard in vitro growth conditions, and the role of Nrg1p in C. auris is currently unknown. Western blot analyses of cell surface and cytosolic proteins of C. auris against anti-CalNrg1 antibody revealed the Nrg1p in both locations. Cell surface localization of Nrg1p in C. auris, an unexpected finding, was further confirmed by immunofluorescence microscopy. Nrg1p expression is uniform across all four clades of C. auris and is dependent on growth conditions. Taken together, the data indicate that C. auris produces several unique proteins during its biofilm growth, which may assist in the skin-colonizing lifestyle of the fungus during its pathogenesis. [ABSTRACT FROM AUTHOR]

Objective: Two-dimensional electrophoresis (2-DE) and MALDI-TOF/TOF mass spectrometry were performed to compare the proteomic alterations of lycorine-treated and control cells to further investigate the anti-multiple myeloma (MM) mechanisms of lycorine. Results: Mass spectrometry results showed that after lycorine treatment of MM cells, 42% of the differentially expressed proteins had subcellular localization, mainly, on mitochondria. Voltage-dependent anion-selective channel protein 2 (VDAC2), the most abundant protein in the outer mitochondrial membrane, was up-regulated after treatment with lycorine and was subsequently verified by western blot analysis. Further studies on mitochondria found that lycorine was able to increase abnormal mitochondria and increase mitochondrial membrane potential. Conclusions: Lycorine can achieve the effect of resisting multiple myeloma by acting on VDAC2 and causing mitochondrial abnormalities. [ABSTRACT FROM AUTHOR]

The proteostasis system of animals, including various types of protein modification during the growth stage, leads to an almost incomprehensible number of possible forms of protein, and each can regulate numerous functions. In the presented work, the composition of muscle tissue protein from different portions of piglets was studied to understand the main muscle protein formation. Comparative analysis of weaned piglets' main muscle protein from l. dorsi, biceps femoris, and brachiocephalicus were analyzed using two-dimensional electrophoresis. Changes in the staining intensity of protein fractions inherent in different muscles were revealed. As part of this work, candidate groups of pig muscle proteins have been selected. Eleven protein spots were revealed for the longest muscle of the back, and seven for the biceps; the muscles of the neck are characterized by indicators of low protein fraction volume. Among the proteins found, myosin light chains, phosphoglycerate mutase, troponins, and adenylate kinase is most likely present. The obtained results of protein identification in muscle tissues, obtained during the intensive growth period, will allow a more detailed understanding of protein regulation, function, and interactions in complex biological systems, which will subsequently be significantly important for biomonitoring health and predicting farm animals productivity. [ABSTRACT FROM AUTHOR]

Abstract Background Trichinella britovi is the second most common species of Trichinella that may affect human health. As an early diagnosis of trichinellosis is crucial for effective treatment, it is important to identify sensitive, specific and common antigens of adult T. britovi worms and muscle larvae. The present study was undertaken to uncover the stage-specific and common proteins of T. britovi that may be used in specific diagnostics. Methods Somatic extracts obtained from two developmental stages, muscle larvae (ML) and adult worms (Ad), were separated using two-dimensional gel electrophoresis (2-DE) coupled with immunoblot analysis. The positively-visualized protein spots specific for each stage were identified through liquid chromatography-tandem mass spectrometry (LC-LC/MS). Results A total of 272 spots were detected in the proteome of T. britovi adult worms (Ad) and 261 in the muscle larvae (ML). The somatic extracts from Ad and ML were specifically recognized by T. britovi-infected swine sera at 10 days post infection (dpi) and 60 dpi, with a total of 70 prominent protein spots. According to immunoblotting patterns and LC-MS/MS results, the immunogenic spots recognized by different pig T. britovi-infected sera were divided into three groups for the two developmental stages: adult stage-specific proteins, muscle larvae stage-specific proteins, and proteins common to both stages. Forty-five Ad proteins (29 Ad-specific and 16 common) and thirteen ML proteins (nine ML-specific and four common) cross-reacted with sera at 10 dpi. Many of the proteins identified in Ad (myosin-4, myosin light chain kinase, paramyosin, intermediate filament protein B, actin-depolymerizing factor 1 and calreticulin) are involved in structural and motor activity. Among the most abundant proteins identified in ML were 14-3-3 protein zeta, actin-5C, ATP synthase subunit d, deoxyribonuclease-2-alpha, poly-cysteine and histide-tailed protein, enolase, V-type proton ATPase catalytic and serine protease 30. Heat-shock protein, intermediate filament protein ifa-1 and intermediate filament protein B were identified in both proteomes. Conclusions To our knowledge, this study represents the first immunoproteomic identification of the antigenic proteins of adult worms and muscle larvae of T. britovi. Our results provide a valuable basis for the development of diagnostic methods. The identification of common components for the two developmental stages of T. britovi may be useful in the preparation of parasitic antigens in recombinant forms for diagnostic use.

Abstract Background Rare data are available on the molecular responses of higher plants to low pH. Seedlings of ‘Sour pummelo’ (Citrus grandis) and ‘Xuegan’ (Citrus sinensis) were treated daily with nutrient solution at a pH of 2.5, 3, or 6 (control) for nine months. Thereafter, we first used 2-dimensional electrophoresis (2-DE) to investigate low pH-responsive proteins in Citrus leaves. Meanwhile, we examined low pH-effects on leaf gas exchange, carbohydrates, ascorbate, dehydroascorbate and malondialdehyde. The objectives were to understand the adaptive mechanisms of Citrus to low pH and to identify the possible candidate proteins for low pH-tolerance. Results Our results demonstrated that Citrus were tolerant to low pH, with a slightly higher low pH-tolerance in the C. sinensis than in the C. grandis. Using 2-DE, we identified more pH 2.5-responsive proteins than pH 3-responsive proteins in leaves. This paper discussed mainly on the pH 2.5-responsive proteins. pH 2.5 decreased the abundances of proteins involved in ribulose bisphosphate carboxylase/oxygenase activation, Calvin cycle, carbon fixation, chlorophyll biosynthesis and electron transport, hence lowering chlorophyll level, electron transport rate and photosynthesis. The higher oxidative damage in the pH 2.5-treated C. grandis leaves might be due to a combination of factors including higher production of reactive oxygen species, more proteins decreased in abundance involved in antioxidation and detoxification, and lower ascorbate level. Protein and amino acid metabolisms were less affected in the C. sinensis leaves than those in the C. grandis leaves when exposed to pH 2.5. The abundances of proteins related to jasmonic acid biosynthesis and signal transduction were increased and decreased in the pH 2.5-treated C. sinensis and C. grandis leaves, respectively. Conclusions This is the first report on low pH-responsive proteins in higher plants. Thus, our results provide some novel information on low pH-toxicity and -tolerance in higher plants.

Abstract Background Azadirachtin, one of the most promising botanical insecticides, has been widely used for pest control. Azadirachtin induces apoptosis in insect cell lines, including Sf9, SL-1 and BTI-Tn-5B1–4. Mitochondrial and lysosomal pathways are likely involved in the azadirachtin-induced apoptosis, however, detailed molecular mechanisms remain largely undefined. Results Azadirachtin-induced apoptosis in Sf9 cells was verified by morphological observation, Hoechst 33258 staining, and a Caspase-3-based analysis. Comparative two-dimensional gel electrophoresis (2-DE) coupled with a linear ion trap quadrupole (LTQ)-MS/MS analysis identified 12 prominent, differentially expressed proteins following azadirachtin treatment. These differentially expressed genes are involved in regulating cytoskeleton development, signal transduction, gene transcription, and cellular metabolism. Knockdown gene expression of a gene encoding a DnaJ homolog enhanced apoptosis induced by azadirachtin in Sf9 cells. Conclusion Azadirachtin treatment induces apoptosis in Sf9 cells and affects expression of multiple genes with functions in cytoskeleton development, signal transduction, gene regulation, and cellular metabolisms. Azadirachtin induces apoptosis at least partially by down-regulation of Sf-DnaJ in Sf9 cells.

Maize rough dwarf disease (MRDD) is a viral disease caused by brown planthopper infestation, and leads to great yield loss, especially in China. Comparative proteomics was performed using maize inbred line Zheng 58 and LN 287. MRDD pathogen was detected as rice black-streaked dwarf virus (RBSDV) by quantitative real time PCR (qRT-PCR) in Shandong Province, China. The modified trichloroacetic acid (TCA)/acetone method was used for soluble protein extraction from leaves. Two-dimensional electrophoresis (2-DE) analysis was performed on 24-cm long, pH 4–7 linear immobilized pH gradient (IPG) strips, and gels were stained with silver and coomassie brilliant blue. We identified 944 proteins expressed in RBSDV infected maize leaves by proteomics approaches. Among these, 44 protein spots that revealed a 1.5-fold difference in intensity were identified by mass spectrometry between mock-inoculated and RBSDV infected samples. Among these, 17 and 26 spots were up-regulated, and 27 and 18 spots were down-regulated in the virus infected samples of Zheng 58 and LN 287, respectively. Differential protein spots were analyzed by mass spectrometry identification, which could be divided into six categories. Furthermore, the expression of stress-related proteins was detected and confirmed by qRT-PCR. This study lays the foundation for further investigations, enabling the enhancement of MRDD resistance in maize.

The interactions between Arabidopsis thaliana and Plutella xylostella have been considered as a model system to unravel the responses of plants to herbivorous insects. Here, we use a 2-DE proteome approach to detect protein expression changes in the leaves of Arabidopsis plants exposed to P. xylostella larval infestation at 27°C within 8 h. Approximately 450 protein spots were reproducibly detected on gels. Of these, comparing healthy and infested leaves, we identified 18 differentially expressed protein spots. Thirteen proteins were successfully identified by MALDI-TOF/MS and LC-ESI-MS/MS. Functional classification analysis indicated that the differentially identified proteins were associated with amino acid, carbohydrate, energy, lipid metabolism, and photosynthesis. In addition, their relative abundances were assessed according to larval pest feeding on Arabidopsis leaves. These data provide valuable new insights for further works in plant-biotic and environmental stress interaction.

Auricularia auricula is one of the most popular edible mushrooms in China. Limited information is available regarding genes and proteins related to A. auricula development from mycelium to mature fruiting body and response to environmental stimuli. With the development of proteomics, this technology platform for addressing this problem is available. A critical step of a good proteome is protein extraction which can gives the best quality and a wide coverage of total proteins. To establish an effective protein extraction protocol suitable for proteomics analysis in mycelium and fruiting body of A. auricula, four previously reported protein extraction protocols including direct extraction, trichloroacetic acid (TCA) precipitation, TCA/acetone precipitation and phenol-based extraction, were compared based on the protein yield and 2-DE patterns. The efficiency of the four methods was evaluated by comparing protein yield and 2-DE patterns. Of the four methods, both TCA precipitation and phenol-based extraction method for A. auricula mycelia protein extraction yielded the good protein separation pattern and higher number of protein spots in 2-DE analysis. Furthermore, phenol-based extraction is superior to the other tested methods for A. auricula fruiting body proteome analysis .

Cattleya tigrina is an endangered Brazilian native orchid with ornamental potential. Biotechnological tools based on tissue culture techniques are effective for its mass propagation and conservation. Protocorm-like bodies (PLBs) were induced from leaves in culture medium supplemented with plant growth regulators, and generated complete in vitro plants. To deepen on the mechanism underlying this morphogenetic route, this study is aimed to identify and characterize differentially expressed proteins during C. tigrina PLBs development. Leaves were inoculated in in culture medium Murashige and Skoog (MS) supplemented with 9 μM Thidiazuron, and PLBs t were collected after 30, 60 and 100 days in culture. Proteomic analysis performed by two-dimensional electrophoresis and MALDI-TOF mass spectrometry detected 122, 132 and 447 protein after 30, 60 and 100 days, respectively. During PLBs development, 36 proteins were differentially expressed. Proteins related to energy and carbohydrate metabolism, cell proliferation, protein processing, and secondary metabolism and phytohormones were assessed showing their relevance in the process of development of PLBs. Many proteins related to metabolic and energy processes were identified after 60 and 100 days in culture, suggesting their role in which is cell division and differentiation. Enolase was only present after 100 days in culture, thus being a candidate as a molecular marker in this developmental phase. [ABSTRACT FROM AUTHOR]

Proteome analysis of embryo and endosperm of Givotia moluccana, a promising angiosperm tree species was carried out using two-dimensional electrophoresis combined with matrix-assisted laser desorption/ionization time of flight to identify differentially abundant proteins and their possible functions. The study revealed the presence of 492 protein spots in the embryo and 432 in the endosperm, and the relative abundance of 44 spots varied significantly between the two. Of the 21 functionally characterized protein spots, 14.3% were involved in metabolism and energy, 28.6% in protein destination and storage, 14.3% in defense and stress and 19% in cell growth and division. The embryo-specific protein ATP-dependent zinc metalloprotease FtsH is predicted to be involved in chloroplast biogenesis and the endosperm-specific protein hydroxyacyl-ACP dehydratase in fatty acid synthesis. These results suggested that the seeds have proteins required for germination and for stress responses. [ABSTRACT FROM AUTHOR]

Glycoproteins are important biomarkers for cancers, while most glycoproteomics biomarkers suffering from low sensitivity and specificity due to their uncharacterized glycan structures. AZGP1 is a potential biomarker for salivary diagnostics of lung cancer, which is used as a model glycoprotein in this study for method development. We initially analyzed salivary N-glycoproteome by using lectin affinity chromatography and more than 300 N-glycoproteins were identified, including AZGP1. 7 gel spots of AZGP1 were resolved by two-dimensional gel electrophoresis and further confirmed by two-dimensional western blot as well as mass spectrometry. The isomeric glycan structures of AZGP1 in these spots were systematically characterized both at composition level and at structure level. Our results revealed 10 glycan compositions for salivary AZGP1, including core fucosylated glycans on Asn128 and sialylated glycans on Asn109 and Asn112. We further compared the glycan structures of salivary AZGP1 from lung cancer group and control group. Accordingly, 14 and 7 potential glycan structures were successfully revealed, respectively. In total, 15 glycan compositions and 22 potential glycan structures were identified and characterized for AZGP1, including some different structures with the same compositions. In particular, 5 potential glycan structures were identified as lung cancer unique signatures. Our developed strategy holds promise for thorough identification of glycan structures on a target glycoprotein biomarker. In-depth characterization of its glycan structures will ultimately enhance its sensitivity and specificity for cancer detection. Image 1 • Affinity chromatography and 2-DE were combined for the separation of glycoproteins. • Robust strategy was developed for glycan identification of target proteomic marker. • 15 glycan compositions and 22 glycan structures were characterized for saliva AZGP1. • Unique glycan structures of saliva AZGP1 were revealed for lung cancer. [ABSTRACT FROM AUTHOR]

We report about the use of protein profiling by two dimensional gel electrophoresis (2-DE) to identify a particular tea clone or cultivar from our recently considered three tea cultivars (TV1, TV6 and HK22/14). Comparison of 2-DE gel pictures for each cultivar provided a unique banding patterns of protein spots on 2-DE gel which remain unchanged after challenging with biotic (Helopeltis and red spider infested) and abiotic (drought) stresses. Therefore, it can be concluded that 2-DE may be used as a protein marker for identifying a desirable tea cultivar to be used in tea breeding programme. [ABSTRACT FROM AUTHOR]

Many pathogenicity factors are involved in the development of primary amoebic meningoencephalitis (PAM) caused by N fowleri. However, most of them are not exclusive for N fowleri and they have not even been described in other nonpathogenic Naegleria species. Therefore, the objective of this work was to identify differential proteins and protein pattern recognition between Naegleria fowleri and Naegleria lovaniensis using antibodies anti‐N fowleri as strategy to find vaccine candidates against meningoencephalitis. Electrophoresis and Western blots conventional and 2‐DE were performed for the identification of antigenic proteins, and these were analysed by the mass spectrometry technique. The results obtained in 2‐DE gels and Western blot showed very notable differences in spot intensity between these two species, specifically those with relative molecular weight of 100, 75, 50 and 19 kDa. Some spots corresponding to these molecular weights were identified as actin fragment, myosin II, heat shock protein, membrane protein Mp2CL5 among others, with differences in theoretical post‐translational modifications. In this work, we found differences in antigenic proteins between both species, proteins that could be used for a further development of vaccines against N fowleri infection. [ABSTRACT FROM AUTHOR]

Global warming has reached an alarming situation, which led to a dangerous climatic condition. The irregular rainfalls and land degradation are the significant consequences of these climatic changes causing a decrease in crop productivity. The effect of drought and its tolerance mechanism, a comparative roots proteomic analysis of chickpea seedlings grown under hydroponic conditions for three weeks, performed at different time points using 2-Dimensional gel electrophoresis (2-DE). After PD-Quest analysis, 110 differentially expressed spots subjected to MALDI-TOF/TOF and 75 spots identified with a significant score. These identified proteins classified into eight categories based on their functional annotation. Proteins involved in carbon and energy metabolism comprised 23% of total identified proteins include mainly glyceraldehyde-3-phosphate dehydrogenase, malate dehydrogenase, transaldolase, and isocitrate dehydrogenase. Proteins related to stress response (heat-shock protein, CS domain protein, and chitinase 2-like) contributed 16% of total protein spots followed by 13% involved in protein metabolism (adenosine kinase 2, and protein disulfide isomerase). ROS metabolism contributed 13% (glutathione S-transferase, ascorbate peroxidase, and thioredoxin), and 9% for signal transduction (actin-101, and 14-3-3-like protein B). Five percent protein identified for secondary metabolism (cinnamoyl-CoA reductase-1 and chalcone-flavononeisomerase 2) and 7% for nitrogen (N) and amino acid metabolism (glutamine synthetase and homocysteine methyltransferase). The abundance of some proteins validated by using Western blotting and Real-Time-PCR. The detailed information for drought-responsive root protein(s) through comparative proteomics analysis can be utilized in the future for genetic improvement programs to develop drought-tolerant chickpea lines. • Comparative proteomics study revealed 75 differentially expressed proteins of chickpea root under drought stress. • Illustrate on many biochemical pathways in stress condition employing the identified proteins. • Up-regulation of the proteins involved in ROS metabolism suggested amelioration of oxidative stress. • Drought also regulated key proteins related to carbon, energy and secondary metabolism, and signalling class. [ABSTRACT FROM AUTHOR]

Adipose derived stem cells (ADSCs) are multipotent and can transdifferentiate into neural stem cells. We investigated the transdifferentiation of ADSCs to neural phenotype (NP) cells using selegiline and two-dimensional electrophoresis (2-DE). The perinephric and inguinal fat of rats was collected and used to isolate ADSCs that were characterized by immunophenotyping using flow cytometry. The ADSCs were differentiated into osteogenic and lipogenic cells. The NP cells were generated using 10−9 mM selegiline and characterized by immunocytochemical staining of nestin and neurofilament 68 (NF-68), and by qRT-PCR of nestin, neurod1 and NF68. Total protein of ADSCs and NP cells was extracted and their proteome patterns were examined using 2-DE. ADSCs carried CD73, CD44 and CD90 cell markers, but not CD34. ADSCs were differentiated into osteocyte and adipocyte lineages. The differentiated NP cells expressed nestin, neuro d1 and NF-68. The proteome pattern of ADSCs was compared with that of NP cells and eight spots showed more than a two fold increase in protein expression. The molecular weights and isoelectric points of these highly expressed proteins were estimated using Melanie software. We compared these results with those of the mouse proteomic database using the protein isoelectric point database, and the functions of the eight proteins in differentiation of NP cells were predicted using the UniProt database. The probable identities of the proteins that showed higher expression in NP cells included cholinesterase, GFRa2, protein kinase C (PKC-eta) and RING finger protein 121. The sequences of the proteins identified from mouse database were aligned by comparing them with similar proteins in rat database using the Basic Local Alignment Search Tool (BLAST). The E values of all aligned proteins were zero, which indicates consistency of the matched protein. These proteins participate in differentiation of the neuron and their overexpression causes ADSCs transdifferentiation into NP cells. [ABSTRACT FROM AUTHOR]

Moringa oleifera Lam is a medicinal plant that had been shown to have anticancer activity. Previously, the hexane fraction of the seeds (HF-CEE) was found to inhibit breast cancer (MCF7) cell proliferation at IC 50 of 130 μg/ml. However, the mechanism by which HF-CEE inhibits MCF7 cells remained unknown. To elucidate the mechanism, we investigated the effect of HF-CEE on apoptosis, cell cycle, and the cell proteome of MCF7 cells. The HF-CEE was prepared by fractionating the non-polar fraction of the ethanolic extract of M. oleifera seeds using the hexane solvent. The antiproliferative effect of HF-CEE on the growth of MDA-MB-231 cells (another breast cancer cells) was performed using Presto Blue assay. The morphologies of the untreated and treated cells (MCF7, MDA-MB-231 and MCF 10A cells) were observed using inverted light microscope. The apoptotic and cell cycle arrest effects of HF-CEE on MCF7 cells were determined using flow cytometry analyses. 2-DE proteomics analysis was employed to identify the protein biomarkers of MCF7 cells that have their expression levels altered by HF-CEE treatment. qRT-PCR method was used to validate the 2-DE results. The gene ontology analysis was carried out using PANTHER server to identify the mechanisms and biological processes that are dependent on the functions of the identified biomarkers. Our results showed that HF-CEE selectively inhibited the proliferation of the breast cancer cells (MCF7 and MDA-MB-231) but not the MCF 10A non-tumor cells. MCF7 and MDA-MB-231 cells treated with HF-CEE showed morphological properties of dead cells, but MCF 10A cells did not. HF-CEE also induced apoptosis and cell cycle arrest at the S and G2/M phases in MCF7 cells. 2-DE analysis revealed that HF-CEE altered the regulation of several cancer-related proteins in MCF7 cells, such as heat shock protein 60 (HSP60), nucleophosmin (NPM), protein disulphide isomerase (PDIA1), phosphoglycerate kinase 1 (PGK1), and reticulocalbin 1 (RCN1). In conclusion, HF-CEE induced cell death in MCF7 cells via apoptosis, cell cycle arrest, and other mechanisms such as the alteration of the expression of proteins that are involved in glycolysis, cancer invasiveness and metastasis among others. Results of this study provide further insights into the mechanisms that underlie the antiproliferative effect of HF-CEE on MCF7 cells. • Moringa oleifera seed hexane fraction (HF-CEE) selectively inhibits breast cancer cells. • HF-CEE induces apoptosis in MCF7 cells. • HF-CEE arrests cell cycle progression at S and G2/M phases. • HF-CEE regulates cancer biomarkers such as HSP60, NPM, PGK1, RCN1, and PDIA1. [ABSTRACT FROM AUTHOR]

Although the economic value of wheat flour is determined by the complement of gluten proteins, these proteins have been challenging to study because of the complexity of the major protein groups and the tremendous sequence diversity among wheat cultivars. The completion of a high-quality wheat genome sequence from the reference wheat Chinese Spring recently facilitated the assembly and annotation of a complete set of gluten protein genes from a single cultivar, making it possible to link individual proteins in the flour to specific gene sequences. In a proteomic analysis of total wheat flour protein from Chinese Spring using quantitative two-dimensional gel electrophoresis combined with tandem mass spectrometry, gliadins or low-molecular-weight glutenin subunits were identified as the predominant proteins in 72 protein spots. Individual spots were associated with 40 of 56 Chinese Spring gene sequences, including 16 of 26 alpha gliadins, 10 of 11 gamma gliadins, six of seven omega gliadins, one of two delta gliadins, and nine of ten LMW-GS. Most genes that were not associated with protein spots were either expressed at low levels in endosperm or encoded proteins with high similarity to other proteins. A wide range of protein accumulation levels were observed and discrepancies between transcript levels and protein levels were noted. This work together with similar studies using other commercial cultivars should provide new insight into the molecular basis of wheat flour quality and allergenic potential. [ABSTRACT FROM AUTHOR]

Background: Both photosynthetic pigments and chloroplasts in plant leaf cells play an important role in deciding on the photosynthetic capacity and efficiency in plants. Systematical investigating the regulatory mechanism of chloroplast development and chlorophyll (Chl) content variation is necessary for clarifying the photosynthesis mechanism for crops. Objective: This study aims to explore the critical regulatory mechanism of leaf color mutation in a yellow–green leaf sesame mutant Siyl-1. Methods: We performed the genetic analysis of the yellow-green leaf color mutation using the F2 population of the mutant Siyl-1. We compared the morphological structure of the chloroplasts, chlorophyll content of the three genotypes of the mutant F2 progeny. We performed the two-dimensional gel electrophoresis (2-DE) and compared the protein expression variation between the mutant progeny and the wild type. Results: Genetic analysis indicated that there were 3 phenotypes of the F2 population of the mutant Siyl-1, i.e., YY type with light-yellow leaf color (lethal); Yy type with yellow-green leaf color, and yy type with normal green leaf color. The yellow-green mutation was controlled by an incompletely dominant nuclear gene, Siyl-1. Compared with the wild genotype, the chloroplast number and the morphological structure in YY and Yy mutant lines varied evidently. The chlorophyll content also significantly decreased (P < 0.05). The 2-DE comparison showed that there were 98 differentially expressed proteins (DEPs) among YY, Yy, and yy lines. All the 98 DEPs were classified into 5 functional groups. Of which 82.7% DEPs proteins belonged to the photosynthesis and energy metabolism group. Conclusion: The results revealed the genetic character of yellow-green leaf color mutant Siyl-1. 98 DEPs were found in YY and Yy mutant compared with the wild genotype. The regulation pathway related with the yellow leaf trait mutation in sesame was analyzed for the first time. The findings supplied the basic theoretical and gene basis for leaf color and chloroplast development mechanism in sesame. [ABSTRACT FROM AUTHOR]