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Sheath rot of rice is one of the most devastating diseases of rice due to its ability to reduce the yield significantly in all rice cultivating areas of India. The purpose of this study was to assess the effectiveness of Bacillus cereus strain RBS-57 against sheath rot disease of rice. In addition, it enables us to understand the molecular mechanism of the host–pathogen-bioagent interactions using a proteomic approach. A combination of seed treatment, seedling dip, and foliar spray with RBS-57 liquid formulation has recorded the lowest sheath rot disease index, both under pot experiment (21.33%) and field conditions (15.33% in trial I and 12.42% in trial II, respectively). In addition to that, RBS-57 application enhanced the plant growth and yield attributes. Moreover, a 2D-PAGE study uses protein profiling to illustrate the molecular response of the tripartite interaction between host–pathogen-bioagent. MALDI-TOF mass spectrometry (MS/MS) analysis identified a total of 20 differentially expressed proteins, primarily implicated in plant metabolism and development, defense response, transcription and signalling. Selected genes were validated by quantitative Real-Time PCR (qRT-PCR) analysis. The alterations in protein abundance and transcripts were positively correlated for all the genes. The present study provides initial insights into the molecular mechanism that underlies the tripartite interaction between the host–pathogen-bioagent in rice plants. [ABSTRACT FROM AUTHOR]

Soil salinity is considered one of the most limiting factors in large agricultural areas, and tomato (Solanum lycopersicum L.) is highly sensitive to this abiotic stress, which exacerbates under drought. The use of halotolerant PGPR is a promising strategy to enhance tomato tolerance and productivity in saline soils. The present study tested the capability of the recently discovered bacterial strain PVr_9 to increase tolerance to salinity in tomato plants in vitro, greenhouse and open field conditions. When inoculated with PVr_9, in vitro seedlings showed a significant increase in primary root length, number of secondary roots and fresh weight both in absence of stress and when exposed to 150 mM NaCl. In plants exposed to salt treatment, PVr_9 upregulated the salt tolerance genes SOS1 and NHX1, and the enzyme prolyl aminopeptidase involved in proline metabolism. Proteins associated with resistance against pathogens were also upregulated by PVr_9 in absence of salt stress. In greenhouse, PVr_9-inoculated tomato plants treated for 1-week with 150 mM NaCl showed a significant increase in shoot fresh biomass, chlorophyll, proline content, and APX activity. Tomato fruits from PVr_9 inoculated plants, exposed in the field to salt stress during the flowering stage, showed higher levels of carotenoids, lycopene, and l-ascorbic acid, as well as an increased antioxidant capacity compared to fruits of uninoculated plants. These results indicate PVr_9 as a potential biostimulant in tomato for better tackling soil salinization in a context of climate change and expansion of coastal salinity, while there is large scope for the improvement of the nutritional characteristics of tomato fruits.

Background and Objectives: Zein proteins of maize endosperm are nutritionally poor, but important for vitreous kernel texture. The Opaque‐2 mutation enhances the protein quality but downregulates zein expression, distorting the kernel texture. Quality protein maize (QPM) is nutritionally improved, hard endosperm maize developed by introgression of endosperm modifiers under an opaque‐2 background. The present study aims to analyze the variability in zein expression patterns in normal, opaque‐2, and QPM lines through sodium dodecyl sulfate‐polyacrylamide gel electrophoresis and two‐dimensional gel electrophoresis analyses. Findings: Results revealed that the total number of zein isoforms is almost similar in opaque‐2, normal, and QPM lines. Overall, in this study, it is concluded that the opaque‐2 mutation specifically affects the expression of high‐molecular‐weight zeins, which are being counterbalanced by the expression of low‐molecular‐weight zein isoforms and nonzein proteins. Conclusion: Genomic data retrieval studies revealed the possible presence of multiple 27 kDa isoforms with varied isoelectric points (pI), which is a subject for further investigation. The differential expression of 15 kDa zein in QPM emerged as a novel player in endosperm modification. Significance and Novelty: The present study provides insight into the diversified expression of zeins in different maize types. The variable isoform expression has the potential to generate stable QPM lines by targeting effective isoforms, ensuring endosperm modification without hampering the nutritional quality. [ABSTRACT FROM AUTHOR]

Abstract Background Low temperature (LT) stress is one of the major environmental stress factors affecting the growth and yield of maize (Zea mays L.). Hence, it is important to unravel the molecular mechanisms behind LT stress tolerance to improve molecular breeding in LT tolerant genotypes. In the present study, two maize genotypes viz. Gurez local from Kashmir Himalaya and tropical grown GM6, were dissected for their LT stress response in terms of accumulation of differentially regulated proteins (DRPs). Leaf proteome analysis at three-leaf stage of maize seedlings subjected to LT stress of 6 °C for a total of 12 h duration was performed using two dimensional gel electrophoresis (2D-PAGE) followed by subsequent identification of the proteins involved. Results After MALDI-TOF (Matrix-assisted laser desorption/ionization-time of flight) and bioinformatics analysis, 19 proteins were successfully identified in Gurez local, while as 10 proteins were found to get successful identification in GM6. The interesting observations from the present investigation is the identification of three novel proteins viz. threonine dehydratase biosynthetic chloroplastic, thylakoidal processing peptidase 1 chloroplastic, and nodulin-like protein, whose role in abiotic stress tolerance, in general, and LT stress, in particular, has not been reported so far. It is important to highlight here that most of LT responsive proteins including the three novel proteins were identified from Gurez local only, owing to its exceptional LT tolerance. From the protein profiles, obtained in both genotypes immediately after LT stress perception, it was inferred that stress responsive protein accumulation and their expression fashion help the Gurez local in seedling establishment and withstand unfavorable conditions as compared to GM6. This was inferred from the findings of pathway enrichment analysis like regulation of seed growth, timing of floral transition, lipid glycosylation, and aspartate family amino acid catabolic processes, besides other key stress defense mechanisms. However, in GM6, metabolic pathways enriched were found to be involved in more general processes including cell cycle DNA replication and regulation of phenylpropanoid metabolism. Furthermore, majority of the qRT-PCR results of the selected proteins demonstrated positive correlation between protein levels and transcript abundance, thereby strengthening our findings. Conclusions In conclusion, our findings reported majority of the identified proteins in Gurez local exhibiting up-regulated pattern under LT stress as compared to GM6. Furthermore, three novel proteins induced by LT stress were found in Gurez local, requiring further functional validation. Therefore, our results offer more insights for elucidating the molecular networks mediating LT stress tolerance in maize.

Background: Low temperature (LT) stress is one of the major environmental stress factors affecting the growth and yield of maize (Zea mays L.). Hence, it is important to unravel the molecular mechanisms behind LT stress tolerance to improve molecular breeding in LT tolerant genotypes. In the present study, two maize genotypes viz. Gurez local from Kashmir Himalaya and tropical grown GM6, were dissected for their LT stress response in terms of accumulation of differentially regulated proteins (DRPs). Leaf proteome analysis at three-leaf stage of maize seedlings subjected to LT stress of 6 °C for a total of 12 h duration was performed using two dimensional gel electrophoresis (2D-PAGE) followed by subsequent identification of the proteins involved. Results: After MALDI-TOF (Matrix-assisted laser desorption/ionization-time of flight) and bioinformatics analysis, 19 proteins were successfully identified in Gurez local, while as 10 proteins were found to get successful identification in GM6. The interesting observations from the present investigation is the identification of three novel proteins viz. threonine dehydratase biosynthetic chloroplastic, thylakoidal processing peptidase 1 chloroplastic, and nodulin-like protein, whose role in abiotic stress tolerance, in general, and LT stress, in particular, has not been reported so far. It is important to highlight here that most of LT responsive proteins including the three novel proteins were identified from Gurez local only, owing to its exceptional LT tolerance. From the protein profiles, obtained in both genotypes immediately after LT stress perception, it was inferred that stress responsive protein accumulation and their expression fashion help the Gurez local in seedling establishment and withstand unfavorable conditions as compared to GM6. This was inferred from the findings of pathway enrichment analysis like regulation of seed growth, timing of floral transition, lipid glycosylation, and aspartate family amino acid catabolic processes, besides other key stress defense mechanisms. However, in GM6, metabolic pathways enriched were found to be involved in more general processes including cell cycle DNA replication and regulation of phenylpropanoid metabolism. Furthermore, majority of the qRT-PCR results of the selected proteins demonstrated positive correlation between protein levels and transcript abundance, thereby strengthening our findings. Conclusions: In conclusion, our findings reported majority of the identified proteins in Gurez local exhibiting up-regulated pattern under LT stress as compared to GM6. Furthermore, three novel proteins induced by LT stress were found in Gurez local, requiring further functional validation. Therefore, our results offer more insights for elucidating the molecular networks mediating LT stress tolerance in maize. [ABSTRACT FROM AUTHOR]

Abstract Background The Australian Imaging and Biomarker Lifestyle (AIBL) study of aging is designed to aid the discovery of biomarkers. The current study aimed to discover differentially expressed plasma proteins that could yield a blood-based screening tool for Alzheimer’s disease. Methods The concentration of proteins in plasma covers a vast range of 12 orders of magnitude. Therefore, to search for medium to low abundant biomarkers and elucidate mechanisms of AD, we immuno-depleted the most abundant plasma proteins and pre-fractionated the remaining proteins by HPLC, prior to two-dimensional gel electrophoresis. The relative levels of approximately 3400 protein species resolved on the 2D gels were compared using in-gel differential analysis with spectrally resolved fluorescent protein detection dyes (Zdyes™). Here we report on analysis of pooled plasma samples from an initial screen of a sex-matched cohort of 72 probable AD patients and 72 healthy controls from the baseline time point of AIBL. Results We report significant changes in variants of apolipoprotein E, haptoglobin, α1 anti-trypsin, inter-α trypsin inhibitor, histidine-rich glycoprotein, and a protein of unknown identity. α1 anti-trypsin and α1 anti-chymotrypsin demonstrated plasma concentrations that were dependent on APOE ε4 allele dose. Our analysis also identified an association with the level of Vitamin D binding protein fragments and complement factor I with sex. We then conducted a preliminary validation study, on unique individual samples compared to the discovery cohort, using a targeted LC-MS/MS assay on a subset of discovered biomarkers. We found that targets that displayed a high degree of isoform specific changes in the 2D gels were not changed in the targeted MS assay which reports on the total level of the biomarker. Conclusions This demonstrates that further development of mass spectrometry assays is needed to capture the isoform complexity that exists in theses biological samples. However, this study indicates that a peripheral protein signature has potential to aid in the characterization of AD.

Proteomics has been widely used to study muscle biology and meat quality traits from different species including beef. Beef proteomics studies allow a better understanding of the biological processes related to meat quality trait determination. This study aimed to decipher by means of two-dimensional electrophoresis (2D-PAGE), mass spectrometry and bioinformatics the changes in post-mortem muscle with a focus on proteins differentially expressed in the Longissimus thoracis (LT) muscle of immunocastrated young heifers and steers. Carcass traits, chemical composition, pH, instrumental color (L*, a*, b*), cooking loss and Warner-Bratzler shear force (WBSF) of meat from F1 Montana-Nellore cattle were also evaluated. Backfat thickness (BFT) and intramuscular fat content (IMF) were 46.8% and 63.6% higher in heifers (p < 0.05), respectively, while evaporation losses (EL) were 10.22% lower compared to steers. No differences (p > 0.05) were observed for tenderness evaluated by WBSF (3, 10, and 17 days post-mortem), pH, and color traits (L*, a* and b*) between the experimental groups. The study revealed several proteins to be differentially expressed proteins in heifers compared steers (p < 0.05). In heifers, proteins involved in nutrient transport (TF, ALB, and MB), energy metabolism (ALDOA, GAPDH, and PKM), and oxidative stress and response to stress (HSPA8 and CA3) were associated with a greater BFT and IMF deposition. The higher expression of these proteins indicated greater oxidative capacity and lower glycolytic activity in the LT muscle of heifers. In steers, there was greater abundance of protein expression related to muscle contraction and proteins of structure (ACTA1, TPM2 and TNNT3), energy metabolism (ENO1, ENO3, PYGM, PGM1 and TPI1) and ATP metabolism (ATP5F1B, PEBP1 and AK1), indicating greater glycogenolysis in LT muscle, suggesting a shift in the glycolytic/oxidative fibers of steers. [ABSTRACT FROM AUTHOR]

Simple Summary: Molecular mechanisms underlying the production of livestock are of great interest at all times. Pituitary adenylate cyclase-activating polypeptide (PACAP) is a multifunctional neuropeptide; it takes part in the regulation of various physiological processes, such as feeding, reproduction, thermoregulation and brain development. This study was conducted to investigate the effect of high plasma PACAP levels and low plasma PACAP levels on the protein composition of blood plasma. Our finding is that plasma PACAP level is associated with the abundance of 10 plasma proteins. The functions of the differentially expressed proteins indicate that the PACAP level of plasma is related with the lipid metabolism and immune status of cattle. Pituitary adenylate cyclase-activating polypeptide (PACAP) is a pleiotropic and multifunctional neuropeptide; it takes part in the regulation of various physiological processes, such as feeding, reproduction, catecholamine synthesis, thermoregulation, motor activity, brain development and neuronal survival. Since PACAP plays important regulatory roles, we hypothesized that the level of PACAP in blood is associated with expression of other proteins, which are involved in different metabolic pathways. The objective of the present study was to compare plasma protein profiles of cows with high and low plasma PACAP levels. Differential proteome analyses were performed by two-dimensional gel electrophoresis (2D-PAGE) followed by tryptic digestion and protein identification by liquid chromatography–mass spectrometry (LC-MS). A total of 210 protein spots were detected, and 16 protein spots showed statistically significant differences (p < 0.05) in the expression levels between groups. Ten spots showed a higher intensity in the high-PACAP-concentration group, while six spots were more abundant in the low-PACAP-concentration group. The functions of the differentially expressed proteins indicate that the PACAP level of plasma is related to the lipid metabolism and immune status of cattle. [ABSTRACT FROM AUTHOR]

There are probably no biological samples that did more to spur interest in proteomics than serum and plasma. The belief was that comparing the proteomes of these samples obtained from healthy and disease-affected individuals would lead to biomarkers that could be used to diagnose conditions such as cancer. While the continuing development of mass spectrometers with greater sensitivity and resolution has been invaluable, the invention of strategic strategies to separate circulatory proteins has been just as critical. Novel and creative separation techniques were required because serum and plasma probably have the greatest dynamic range of protein concentration of any biological sample. The concentrations of circulating proteins can range over twelve orders of magnitude, making it a challenge to identify low-abundance proteins where the bulk of the useful biomarkers are believed to exist. The major goals of this article are to (i) provide an historical perspective on the rapid development of serum and plasma proteomics; (ii) describe various separation techniques that have made obtaining an in-depth view of the proteome of these biological samples possible; and (iii) describe applications where serum and plasma proteomics have been employed to discover potential biomarkers for pathological conditions. [ABSTRACT FROM AUTHOR]

Plants have evolved diverse mechanisms to cope with damages caused by insect herbivores. Proteomics can greatly contribute to the understanding of plant changes in response to herbivory. Here, two-dimensional polyacrylamide gel electrophoresis was used in a comparative proteomics approach to survey responses of pistachio seedlings to herbivory of the nymphal stages of the common pistachio psylla. Protein spots (~ 360) were reproducibly detected on each gel that led to the identification of 24 differentially expressed proteins via nano liquid chromatography tandem mass spectrometry (nLC-MS/MS). The identified proteins were involved in photosynthesis, glycolysis, signaling, protein folding, homeostasis, apoptotic processes, and stress response. Pyrophosphatase 6, controlled tumor protein, ferritin1, succinate dehydrogenase 1–1 were the proteins reported for the first time in response to herbivory. It seems that increased growth rate is a major strategy against herbivory stress of the psylla. The results of this study provide valuable information for understanding plants—herbivores interaction. [ABSTRACT FROM AUTHOR]

In the current study, some proteomic methods containing 2D-PAGE and FT-IR techniques were performed to screen gliadin, the subunit of gluten protein, and transformation between a and ß sheet forms of this protein was evaluated. The protein concentration of the samples varied between flour types and also cooked form of these samples. We focused on Amide A, Amide I and phosphorylated protein regions on the spectrums achieved by FT-IR. Gliadin structure was dramatically differed when the raw material was formed in baked form. While Amide A vibration which is related to N-H streching increased for the bread form of the white flour, Amide I which is related to C=O streching decreased when the raw material changed in the cooked form. It can be concluded that the type of flour used in bread production and the type of baking were effective on gluten structure and amount of the final product. [ABSTRACT FROM AUTHOR]

Background: The Australian Imaging and Biomarker Lifestyle (AIBL) study of aging is designed to aid the discovery of biomarkers. The current study aimed to discover differentially expressed plasma proteins that could yield a blood-based screening tool for Alzheimer's disease. Methods: The concentration of proteins in plasma covers a vast range of 12 orders of magnitude. Therefore, to search for medium to low abundant biomarkers and elucidate mechanisms of AD, we immuno-depleted the most abundant plasma proteins and pre-fractionated the remaining proteins by HPLC, prior to two-dimensional gel electrophoresis. The relative levels of approximately 3400 protein species resolved on the 2D gels were compared using in-gel differential analysis with spectrally resolved fluorescent protein detection dyes (Zdyes™). Here we report on analysis of pooled plasma samples from an initial screen of a sex-matched cohort of 72 probable AD patients and 72 healthy controls from the baseline time point of AIBL. Results: We report significant changes in variants of apolipoprotein E, haptoglobin, α1 anti-trypsin, inter-α trypsin inhibitor, histidine-rich glycoprotein, and a protein of unknown identity. α1 anti-trypsin and α1 anti-chymotrypsin demonstrated plasma concentrations that were dependent on APOE ε4 allele dose. Our analysis also identified an association with the level of Vitamin D binding protein fragments and complement factor I with sex. We then conducted a preliminary validation study, on unique individual samples compared to the discovery cohort, using a targeted LC-MS/MS assay on a subset of discovered biomarkers. We found that targets that displayed a high degree of isoform specific changes in the 2D gels were not changed in the targeted MS assay which reports on the total level of the biomarker. Conclusions: This demonstrates that further development of mass spectrometry assays is needed to capture the isoform complexity that exists in theses biological samples. However, this study indicates that a peripheral protein signature has potential to aid in the characterization of AD. [ABSTRACT FROM AUTHOR]

Introduction: Infective endocarditis (IE) has high mortality, partly due to delayed diagnosis. No biomarker can identify IE in patients with fever and clinical picture of infection. To find putative biomarkers we analyzed serum levels of two proteins found in cardiac valves, fibulin-1 (n = 696) and osteoprotegerin (n = 689) among patients on clinical suspicion of IE. Proteomic analyses were performed in 24 patients with bacteremia, 12 patients with definite IE and 12 patients with excluded IE. Methods: Fibulin-1 and osteoprotegerin were studied by enzyme linked immunosorbent assay (ELISA). Proteomic analyses were conducted by 2-dimensional polyacrylamid gel electrophoresis (2D-PAGE) and label-free quantitative liquid chromatography - tandem mass spectrometry (LFQ LC-MS/MS). Controls for 2D 2D-PAGE and LFQ LC-MS/MS had bacteremia and excluded IE. Results: Osteoprotegerin levels were significantly increased in IE patients compared with non-IE patients. Fibulin-1 showed no difference. 2D-PAGE showed significant differences of 6 proteoforms: haptoglobin, haptoglobin-related protein, α-2-macroglobulin, apolipoprotein A-I and ficolin-3. LFQ LC-MS/MS analysis revealed significant level changes of 7 proteins: apolipoprotein L1, complement C1q subcomponent B and C, leukocyte immunoglobulin-like receptor subfamily A member 3, neuropilin-2, multimerin-1 and adiponectin. Conclusions: The concentration changes in a set of proteoforms/proteins suggest that stress and inflammation responses are perturbed in patients with IE compared to patients with bacteremia without IE.

Over the last two decades, a large number of non-communicable/chronic disorders reached an epidemic level on a global scale such as diabetes mellitus type 2, cardio-vascular disease, several types of malignancies, neurological and eye pathologies—all exerted system's enormous socio-economic burden to primary, secondary, and tertiary healthcare. The paradigm change from reactive to predictive, preventive, and personalized medicine (3PM/PPPM) has been declared as an essential transformation of the overall healthcare approach to benefit the patient and society at large. To this end, specific biomarker panels are instrumental for a cost-effective predictive approach of individualized prevention and treatments tailored to the person. The source of biomarkers is crucial for specificity and reliability of diagnostic tests and treatment targets. Furthermore, any diagnostic approach preferentially should be noninvasive to increase availability of the biomaterial, and to decrease risks of potential complications as well as concomitant costs. These requirements are clearly fulfilled by tear fluid, which represents a precious source of biomarker panels. The well-justified principle of a "sick eye in a sick body" makes comprehensive tear fluid biomarker profiling highly relevant not only for diagnostics of eye pathologies but also for prediction, prognosis, and treatment monitoring of systemic diseases. One prominent example is the Sicca syndrome linked to a cascade of severe complications that include dry eye, neurologic, and oncologic diseases. In this review, protein profiles in tear fluid are highlighted and corresponding biomarkers are exemplified for several relevant pathologies, including dry eye disease, diabetic retinopathy, cancers, and neurological disorders. Corresponding analytical approaches such as sample pre-processing, differential proteomics, electrophoretic techniques, high-performance liquid chromatography (HPLC), enzyme-linked immuno-sorbent assay (ELISA), microarrays, and mass spectrometry (MS) methodology are detailed. Consequently, we proposed the overall strategies based on the tear fluid biomarkers application for 3P medicine practice. In the context of 3P medicine, tear fluid analytical pathways are considered to predict disease development, to target preventive measures, and to create treatment algorithms tailored to individual patient profiles. [ABSTRACT FROM AUTHOR]

To study the mechanism of heat-induced protein aggregates, skim milk was heated at 55, 65, 75, 85, and 95°C for 30 s. Then, the sulfhydryl content, surface hydrophobicity, and secondary structure of heat-treated skim milk were studied. Treating skim milk at different temperatures induced a decrease in sulfhydryl content (75.9% at 95°C) and an increase in surface hydrophobicity (44% at 95°C) with a disrupted secondary structure containing random coil, β-sheet, and β-turn of skim milk proteins. The change in these properties facilitated aggregate formation through disulfide bonds and hydrophobicity interaction. Microstructural observation also showed a higher degree of aggregation when skim milk was heated at 85 and 95°C. The result of two-dimensional polyacrylamide gel electrophoresis demonstrated that the aggregates consisted of a high proportion of κ-casein, β-lactoglobulin, and other whey proteins.

Abstract Background Maize is one of the important cereal food crops in the world. High temperature stress causes adverse influence on plant growth. When plants are exposed to high temperatures, they produce heat shock proteins (HSPs), which may impart a generalized role in tolerance to heat stress. Proteome analysis was performed in plant to assess the changes in protein types and their expression levels under abiotic stress. The purpose of the study is to explore which proteins are involved in the response of the maize plant to heat shock treatment. Results We investigated the responses of abundant proteins of maize leaves, in an Egyptian inbred line of maize “K1”, upon heat stress through two-dimensional electrophoresis (2-DE) on samples of maize leaf proteome. 2-DE technique was used to recognize heat-responsive protein spots using Coomassie Brilliant Blue (CBB) and silver staining. In 2-D analysis of proteins from plants treated at 45 °C for 2 h, the results manifested 59 protein spots (4.3%) which were reproducibly detected as new spots where did not present in the control. In 2D for treated plants for 4 h, 104 protein spots (7.7%) were expressed only under heat stress. Quantification of spot intensities derived from heat treatment showed that twenty protein spots revealed clear differences between the control and the two heat treatments. Nine spots appeared with more intensity after heat treatments than the control, while four spots appeared only after heat treatments. Five spots were clearly induced after heat treatment either at 2 h or 4 h and were chosen for more analysis by LC-MSMS. They were identified as ATPase beta subunit, HSP26, HSP16.9, and unknown HSP/Chaperonin. Conclusion The results revealed that the expressive level of the four heat shock proteins that were detected in this study plays important roles to avoid heat stress in maize plants.

Recent weather fluctuations imposing heat stress at the time of wheat grain filling cause frequent losses in grain yield and quality. Field-based studies for understanding the effect of terminal heat stress on wheat are complicated by the effect of multiple confounding variables. In the present study, the effect of day and day–night combined heat stresses during the grain-filling stage was studied using gene expression and proteomics approaches. The gene expression analysis was performed by using real-time quantitative PCR (RT-qPCR). The expression of genes related to the starch biosynthetic pathway, starch transporters, transcription factors, and stress-responsive and storage proteins, at four different grain developmental stages, indicated the involvement of multiple pathways. Under the controlled conditions, their expression was observed until 28 days after anthesis (DAA). However, under the day stress and day–night stress, the expression of genes was initiated earlier and was observed until 14 DAA and 7 DAA, respectively. The protein profiles generated using two-dimensional polyacrylamide gel electrophoresis (2D-PAGE) and matrix-assisted laser desorption/ionization time-of-flight mass spectroscopy (MALDI-TOF MS/MS) showed a differential expression of the proteins belonging to multiple pathways that included the upregulation of proteins related to the translation, gliadins, and low-molecular-weight (LMW) glutenins and the downregulation of proteins related to the glycolysis, photosynthesis, defense, and high-molecular-weight (HMW) glutenins. Overall, the defense response to the day heat stress caused early gene expression and day–night heat stress caused suppression of gene expression by activating multiple pathways, which ultimately led to the reduction in grain-filling duration, grain weight, yield, and processing quality.

Recent weather fluctuations imposing heat stress at the time of wheat grain filling cause frequent losses in grain yield and quality. Field-based studies for understanding the effect of terminal heat stress on wheat are complicated by the effect of multiple confounding variables. In the present study, the effect of day and day–night combined heat stresses during the grain-filling stage was studied using gene expression and proteomics approaches. The gene expression analysis was performed by using real-time quantitative PCR (RT-qPCR). The expression of genes related to the starch biosynthetic pathway, starch transporters, transcription factors, and stress-responsive and storage proteins, at four different grain developmental stages, indicated the involvement of multiple pathways. Under the controlled conditions, their expression was observed until 28 days after anthesis (DAA). However, under the day stress and day–night stress, the expression of genes was initiated earlier and was observed until 14 DAA and 7 DAA, respectively. The protein profiles generated using two-dimensional polyacrylamide gel electrophoresis (2D-PAGE) and matrix-assisted laser desorption/ionization time-of-flight mass spectroscopy (MALDI-TOF MS/MS) showed a differential expression of the proteins belonging to multiple pathways that included the upregulation of proteins related to the translation, gliadins, and low-molecular-weight (LMW) glutenins and the downregulation of proteins related to the glycolysis, photosynthesis, defense, and high-molecular-weight (HMW) glutenins. Overall, the defense response to the day heat stress caused early gene expression and day–night heat stress caused suppression of gene expression by activating multiple pathways, which ultimately led to the reduction in grain-filling duration, grain weight, yield, and processing quality. [ABSTRACT FROM AUTHOR]

A proteomic method combining two-dimensional polyacrylamide gel electrophoresis (2D-PAGE) and matrix-assisted laser desorption/ionization-time-of-flight/time-of-flight mass spectrometry (MALDI-TOF/TOF) was used to compare the protein expression profiles of phosphine-resistant (PH3-R) and -susceptible (PH3-S) strains of Sitophilus oryzae. Thirty-nine differentially expressed protein spots were identified between the PH3-R and PH3-S strains; 20 protein spots were upregulated, and 19 protein spots were downregulated in the PH3-R strain compared with their expression in the PH3-S strain. In particular, cytochrome oxidase subunit I showed 15-fold higher expression in the PH3-R strain than in the PH3-S strain. Additionally, citrate synthase 2, delta-1-pyrolline-5-carboxylate dehydrogenase, and triose-phosphate isomerase were highly expressed in the PH3-R strain. In summary, our study has improved understanding of the molecular mechanisms of phosphine resistance in the rice weevil. [ABSTRACT FROM AUTHOR]

Pituitary adenylate cyclase-activating polypeptide (PACAP) is a pleiotropic and multifunctional neuropeptide; it takes part in the regulation of various physiological processes, such as feeding, reproduction, catecholamine synthesis, thermoregulation, motor activity, brain development and neuronal survival. Since PACAP plays important regulatory roles, we hypothesized that the level of PACAP in blood is associated with expression of other proteins, which are involved in different metabolic pathways. The objective of the present study was to compare plasma protein profiles of cows with high and low plasma PACAP levels. Differential proteome analyses were performed by two-dimensional gel electrophoresis (2D-PAGE) followed by tryptic digestion and protein identification by liquid chromatography–mass spectrometry (LC-MS). A total of 210 protein spots were detected, and 16 protein spots showed statistically significant differences (p < 0.05) in the expression levels between groups. Ten spots showed a higher intensity in the high-PACAP-concentration group, while six spots were more abundant in the low-PACAP-concentration group. The functions of the differentially expressed proteins indicate that the PACAP level of plasma is related to the lipid metabolism and immune status of cattle.

There are probably no biological samples that did more to spur interest in proteomics than serum and plasma. The belief was that comparing the proteomes of these samples obtained from healthy and disease-affected individuals would lead to biomarkers that could be used to diagnose conditions such as cancer. While the continuing development of mass spectrometers with greater sensitivity and resolution has been invaluable, the invention of strategic strategies to separate circulatory proteins has been just as critical. Novel and creative separation techniques were required because serum and plasma probably have the greatest dynamic range of protein concentration of any biological sample. The concentrations of circulating proteins can range over twelve orders of magnitude, making it a challenge to identify low-abundance proteins where the bulk of the useful biomarkers are believed to exist. The major goals of this article are to (i) provide an historical perspective on the rapid development of serum and plasma proteomics; (ii) describe various separation techniques that have made obtaining an in-depth view of the proteome of these biological samples possible; and (iii) describe applications where serum and plasma proteomics have been employed to discover potential biomarkers for pathological conditions.

The differences in tolerance to high temperatures were investigated on the basis of gene expressions in two strawberry (Fragaria x ananassa Duch) cultivars which were previously determined as high temperature tolerant (Redlands Hope = R. Hope) and sensitive (Festival). Plants were exposed incrementally to 35, 40, 45, and finally 50 °C for 24 h. qRT-PCR analyses were carried out with 19 known sequences from the databases. Protein expression analyses were based on SDS-PAGE results, sequenced and then separated due to their isoelectric points. Expression levels were determined at 35, 40, and 45 °C. According to the results, tolerance of 'R. Hope' to high temperature stress can be explained with the coordination of Hsp70, Hsp90, and small heat shock proteins (sHsps) having a vital and supplementary role in stress response. Sensitive cultivar 'Festival' can respond to high temperatures only with the low molecular weight protein and transcripts that do not take a central role in high temperature stress response. Moreover, allergen gene expression triggered by high temperature were detected in both cultivars with different expression levels. The greater expression level in allergen genes observed in the sensitive cultivar 'Festival' under high temperature indicates that there is possibly a negative correlation between expression level in allergen genes and heat stress tolerance. Future studies addressing allergen gene expression under high temperature stress are required to confirm on these findings and to expand on the potential use as a molecular marker in breeding process for enhanced tolerance to high temperature. [ABSTRACT FROM AUTHOR]

• Biological markers are missing for infective endocarditis. • Most studies miss a proper control group – bacteremia without endocarditis. • Proteomic analyses identified a set of potential diagnostic markers. Infective endocarditis (IE) has high mortality, partly due to delayed diagnosis. No biomarker can identify IE in patients with fever and clinical picture of infection. To find putative biomarkers we analyzed serum levels of two proteins found in cardiac valves, fibulin-1 (n = 696) and osteoprotegerin (n = 689) among patients on clinical suspicion of IE. Proteomic analyses were performed in 24 patients with bacteremia, 12 patients with definite IE and 12 patients with excluded IE. Fibulin-1 and osteoprotegerin were studied by enzyme linked immunosorbent assay (ELISA). Proteomic analyses were conducted by 2-dimensional polyacrylamid gel electrophoresis (2D-PAGE) and label-free quantitative liquid chromatography - tandem mass spectrometry (LFQ LC-MS/MS). Controls for 2D 2D-PAGE and LFQ LC-MS/MS had bacteremia and excluded IE. Osteoprotegerin levels were significantly increased in IE patients compared with non-IE patients. Fibulin-1 showed no difference. 2D-PAGE showed significant differences of 6 proteoforms: haptoglobin, haptoglobin-related protein, α-2-macroglobulin, apolipoprotein A-I and ficolin-3. LFQ LC-MS/MS analysis revealed significant level changes of 7 proteins: apolipoprotein L1, complement C1q subcomponent B and C, leukocyte immunoglobulin-like receptor subfamily A member 3, neuropilin-2, multimerin-1 and adiponectin. The concentration changes in a set of proteoforms/proteins suggest that stress and inflammation responses are perturbed in patients with IE compared to patients with bacteremia without IE. [ABSTRACT FROM AUTHOR]

Haemocytes play a dominant role in shellfish immunity, being considered the main defence effector cells in molluscs. These cells are known to be responsible for many functions, including chemotaxis, cellular recognition, attachment, aggregation, shell repair and nutrient transport and digestion. There are two basic cell types of bivalve haemocytes morphologically distinguishable, hyalinocytes and granulocytes; however, functional differences and specific abilities are poorly understood: granulocytes are believed to be more efficient in killing microorganisms, while hyalinocytes are thought to be more specialised in clotting and wound healing. A proteomic approach was implemented to find qualitative differences in the protein profile between granulocytes and hyalinocytes of the European flat oyster, Ostrea edulis , as a way to evaluate functional differences. Oyster haemolymph cells were differentially separated by Percoll® density gradient centrifugation. Granulocyte and hyalinocyte proteins were separated by 2D-PAGE and their protein profiles were analysed and compared with PD Quest software; the protein spots exclusive for each haemocyte type were excised from gels and analysed by MALDI-TOF/TOF with a combination of mass spectrometry (MS) and MS/MS for sequencing and protein identification. A total of 34 proteins were identified, 20 unique to granulocytes and 14 to hyalinocytes. The results suggested differences between the haemocyte types in signal transduction, apoptosis, oxidation reduction processes, cytoskeleton, phagocytosis and pathogen recognition. These results contribute to identify differential roles of each haemocyte type and to better understand the oyster immunity mechanisms, which should help to fight oyster diseases. Image 1 • A proteomic approach was used to study differences between oyster haemocyte types. • The number of separated proteins was higher in granulocytes than in hyalinocytes. • Proteins unique to each cell type were identified suggesting functional differences. • Proteins involved in apoptosis and intracellular digestion corresponded to granulocytes. • Proteins involved in shell production and wound healing corresponded to hyalinocytes. [ABSTRACT FROM AUTHOR]

Plastid casein kinase 2 (CK2), which is a major Ser/Thr‐specific enzyme in higher organisms, plays an essential role in plant development and diverse abiotic stresses. CKB1 is a regulatory subunit beta of CK2. To expand our understand of functions of the CKB1 gene in Arabidopsis thaliana, protein changes among wild‐type (WT) and CKB1 gain‐ and loss‐of‐function mutants were compared.Proteins extracted from the CKB1 knockout mutant and overexpressing mutant were compared with Col‐0 plants using 2D‐PAGE. Proteins regulated by CKB1 were identified with matrix‐assisted laser desorption ionisation time‐of‐flight/time‐of‐flight mass spectrometry (MALDI‐TOF/TOF), and its transcript was verified by qRT‐PCR. Bioinformatics analysis, including gene ontology and protein–protein interaction analysis, were employed.The results of mass spectra and bioinformatics analysis suggest that CKB1 may have functions in regulation of the ribosomal protein L10 (RPL10) family and is involved in ultraviolet‐B (UV‐B) response. Furthermore, qRT‐PCR verification showed CKB1 expression was up‐regulated by UV‐B stress. The expression levels of five genes in the RPL10 family were reduced in the ckb1 T‐DNA insertion mutants, whereas they increased in the CKB1 overexpressing mutants under both normal conditions and UV‐B treatment.In conclusion, CKB1 has important functions in UV‐B radiation stress. Our study implies that CKB1 positively regulates UV‐B radiation stress signalling, possibly through modulating expression of the RPL10 family. [ABSTRACT FROM AUTHOR]

Proteomics approach was used mainly in the samples for microorganisms and animal tissues until early 2000s, the period during which the plant proteomics studies were scarcely available. Tough nature of cell membranes, existence of complex and diverse range of secondary metabolites, high abundance of pigments, proteases, polyphenols, polysaccharides and lipids caused a number of problems for preparation and resolution of the protein samples. However, proteomics approach in plant area had become widespread with the help of continuous efforts in order to overcome these difficulties. This review is prepared comprehensively from the preparation of the samples to the identification of the proteins and it aims to serve as a detailed source for the researchers interested in plant proteomics, especially for young investigators. [ABSTRACT FROM AUTHOR]

A proteomic method combining two-dimensional polyacrylamide gel electrophoresis (2D-PAGE) and matrix-assisted laser desorption/ionization-time-of-flight/time-of-flight mass spectrometry (MALDI-TOF/TOF) was used to compare the protein expression profiles of phosphine-resistant (PH3-R) and -susceptible (PH3-S) strains of Sitophilus oryzae. Thirty-nine differentially expressed protein spots were identified between the PH3-R and PH3-S strains; 20 protein spots were upregulated, and 19 protein spots were downregulated in the PH3-R strain compared with their expression in the PH3-S strain. In particular, cytochrome oxidase subunit I showed 15-fold higher expression in the PH3-R strain than in the PH3-S strain. Additionally, citrate synthase 2, delta-1-pyrolline-5-carboxylate dehydrogenase, and triose-phosphate isomerase were highly expressed in the PH3-R strain. In summary, our study has improved understanding of the molecular mechanisms of phosphine resistance in the rice weevil.

Background: Maize is one of the important cereal food crops in the world. High temperature stress causes adverse influence on plant growth. When plants are exposed to high temperatures, they produce heat shock proteins (HSPs), which may impart a generalized role in tolerance to heat stress. Proteome analysis was performed in plant to assess the changes in protein types and their expression levels under abiotic stress. The purpose of the study is to explore which proteins are involved in the response of the maize plant to heat shock treatment. Results: We investigated the responses of abundant proteins of maize leaves, in an Egyptian inbred line of maize "K1", upon heat stress through two-dimensional electrophoresis (2-DE) on samples of maize leaf proteome. 2-DE technique was used to recognize heat-responsive protein spots using Coomassie Brilliant Blue (CBB) and silver staining. In 2-D analysis of proteins from plants treated at 45 °C for 2 h, the results manifested 59 protein spots (4.3%) which were reproducibly detected as new spots where did not present in the control. In 2D for treated plants for 4 h, 104 protein spots (7.7%) were expressed only under heat stress. Quantification of spot intensities derived from heat treatment showed that twenty protein spots revealed clear differences between the control and the two heat treatments. Nine spots appeared with more intensity after heat treatments than the control, while four spots appeared only after heat treatments. Five spots were clearly induced after heat treatment either at 2 h or 4 h and were chosen for more analysis by LC-MSMS. They were identified as ATPase beta subunit, HSP26, HSP16.9, and unknown HSP/Chaperonin. Conclusion: The results revealed that the expressive level of the four heat shock proteins that were detected in this study plays important roles to avoid heat stress in maize plants. [ABSTRACT FROM AUTHOR]

Seed longevity is a very important characteristic controlling seed quality. However, the mechanism underlying this characteristic is poorly understood. In this study, 'FH7185', a storable rice variety, the germination rate of which was 66.63%, much higher than the control after artificial ageing under relative humidity 88% and 42°C for 21 days. Different methods were applied to reveal the involvement of proteins during ageing of rice seeds. In total, 35 differential proteins were identified from 2D‐PAGE, and 3,719 proteins from iTRAQ analysis. A comparison of these two methods showed that not all protein types could be detected on the 2D‐PAGE gels, and the dynamic range was somewhat limited. So the comprehensive proteome map from the iTRAQ analysis was used to identify the quantitatively regulated proteins that played roles in seed longevity, and found that the most marked change was the increased abundance of many metabolic enzymes, especially the ones involved in α‐linolenic acid metabolism in the embryos during ageing. OsLOX2 and OsLOX3 had a negative effect on seed longevity, which were lower in FH7185 than the control. The dehydrogenase (LOC_Os07g44430) which played a major catalytic role in lipid peroxidation, also changed significantly during ageing. Therefore, OsLOX2, OsLOX3 and Loc_OS 07G44430 may be involved in the regulation of seed longevity with a synergistic effect. [ABSTRACT FROM AUTHOR]

Plant growth-promoting rhizobacteria such as Azospirillum brasilense Sp7 can protect plants against viruses but the molecular basis of this phenomenon is unclear. We therefore used differential proteomics to study two pathosystems in the presence and absence of Sp7 during early vegetative growth: tomato (Solanum lycopersicum L. cv. Boludo)/Potato virus X (PVX, KJ631111)/Sp7, and maize (Zea mays cv. B73)/Maize dwarf mosaic virus (MDMV, AM110558)/Sp7). In the maize/MDMV system, PDQuest revealed significant variations in the levels of 19 proteins compared to uninfected controls, including the upregulation of NADP-dependent malic enzyme as a form of host-specific viral anticipation, causing a simultaneous increase in the abundance of proteins related to photosynthesis and plastid functions. However, 42 proteins varied significantly in the maize/MDMV/Sp7 system, including the upregulation of radical-scavenging enzymes and proteins related to methionine metabolism, the glutathione-ascorbate cycle and photosynthesis, increasing the photosynthetic rate. In the tomato/PVX system, we observed significant variations in the levels of 58 proteins reflecting the disruption of the Calvin-Benson cycle, responses to oxidative stress and the inhibition of photosystem II (PSII) activity. We identified 26 proteins that varied in the tomato/PVX/Sp7 system; PSII and plastid proteins transiently declined but partially recovered over time as the Calvin-Benson cycle was induced to compensate. Sp7 therefore triggers induced systemic resistance in both pathosystems without affecting the virus titer, although it does delay the appearance of MDMV. The role of ribulose-1.5-bisphosphate carboxylase/oxygenase small subunit as a host target for viruses is discussed in both pathosystems. [ABSTRACT FROM AUTHOR]

Protocols that improve growth performance in fish while assuring product quality are important for aquaculture. Fasting followed by refeeding may promote compensatory growth, thus optimizing growth performance. During fasting and refeeding, fast-twitch muscle, which comprises most of fish fillet, undergoes intense plasticity. In this work, we studied the proteome of pacu (Piaractus mesopotamicus) fast-twitch muscle after 30 days of fasting (D30), 30 days of refeeding (D60) and 60 days of refeeding (D90) with two-dimensional electrophoresis, mass spectrometry and bioinformatics. Body mass, growth rate and muscle histology were also assessed. At D30, fish presented muscle catabolism and decreased growth. Proteomic analysis showed that metabolism proteins were the most affected, up and downregulated. Cytoskeleton and amino acid biosynthesis proteins were downregulated, while nuclear and regulatory proteins were upregulated. At D60, fish showed accelerated growth, despite the body mass not completely recovering. Metabolism proteins were still the most affected. Amino acid biosynthesis proteins became upregulated, while cytoskeleton proteins remained downregulated. At D90, the fish presented total compensatory growth. Many metabolic proteins were up or downregulated. Few cytoskeleton proteins remained differentially expressed. Amino acid biosynthesis proteins were mostly upregulated, but less than at D60. Prolonged fasting followed by refeeding also led to the regulation of possible meat quality biomarkers, such as antioxidant enzymes. This fact suggests possible consequences of this protocol on fish meat quality. Our work also enriches our knowledge on proteomic changes during muscle plasticity that occur during fasting and refeeding diet protocols. Unlabelled Image • 30 days of fasting and 60 days of refeeding led to total compensatory growth in pacu. • The first proteomic analysis of skeletal muscle in fasted-refed fish was performed. • Prolonged fasting and refeeding may affect pacu meat quality. [ABSTRACT FROM AUTHOR]

A proteomic method combining two-dimensional polyacrylamide gel electrophoresis and tandem mass spectrometry was used to compare the hemolymph expression profiles of a beta-cypermethrin-resistant Blattella germanica L. strain (R) and a susceptible strain (S) after 24 h of beta-cypermethrin induction. The results showed that there were 42 differentially expressed proteins after induction of the R strain: 4 proteins were upregulated and 38 proteins were downregulated. One hundred one hemolymph proteins were differentially expressed after induction of the S strain: 53 proteins were upregulated and 48 proteins were downregulated. The identified proteins were mainly classified into the following categories: energy metabolism proteins such as arginine kinase and triose phosphate isomerase, detoxification-related proteins such as glutathione S-transferases (GSTs), signal molecule-regulated proteins such as nitric oxide synthase (NOS), and other proteins such as kinetic-related proteins and gene expression–related proteins. Several proteins show significant differences in response to short-term stress and long-term adaptation, and differential expression of these proteins reflects an overall change in cellular structure and metabolism associated with resistance to pyrethroid insecticides. In summary, our research has improved the understanding of the molecular mechanisms of beta-cypermethrin resistance in German cockroaches, which will facilitate the development of rational methods to improve the management of this pest. [ABSTRACT FROM AUTHOR]

Abstract The pathogenic bacteria Photobacterium damselae subsp. piscicida affects the development of Solea senegalensis culture. Vaccines made with inactivated cells have produced a relative protection against the sickness, however the administration of subcellular and purified antigens as vaccine could increase the effectiveness of the immune response. Thus, the aim of this work was the determination of antigens of P. damselae subsp. piscicida involved in the specific immune response of S. senegalensis. Fish were immunized by intraperitoneal injection (i.p.) with inactivated extracellular polymeric substances (ECP) and whole cells of P. damselae subsp. piscicida , and Freund's incomplete adjuvant. Two months later fish were boosted with the same antigens. Serum from fish was collected to determine by ELISA the title of antibodies against subcellular fractions of bacteria (ECP, capsule, outer membrane proteins, O antigen and formalized whole cells). Significant differences were found between control and immunized fish, but differences between first immunization and booster were only found for O antigen and capsule. Western blots derived from 2D-PAGE of ECP and Outer Membrane Proteins (OMP), using sole immunized serum, detected two high reactive antigens from ECP. Proteins were identified, by mass spectrometry, as ATP-dependent metalloprotease and Telurite resistance proteins. In the case of OMP, three antigenic proteins were detected and identified as Nrfa Y218f, Anti-oxidant AhpC/TSA, and a protein domain DNA binding heat shock related. Highlights • Sole specimens were immunized with inactivated cells of P. damselae subsp. piscicida and ECP to obtain antiserum. • Specific immune response was detected by ELISA using P. damselae subsp. piscicida antigens. • 2D Western blots of ECP and OMP, using sole immunized serum, identified five immunogenic proteins. • Two reactive antigens were identified from ECP: ATP-dependent metalloprotease and Telurite resistance proteins. • In OMP were detected three proteins: Nrfa Y218f, Anti-oxidant AhpC/TSA, and a protein domain DNA binding heat shock related. [ABSTRACT FROM AUTHOR]

In previous work, two independent Arabidopsis thaliana Ac/Ds transposon insertional mutant lines, atnp01 and atnp02, were identified that showed a higher level of tolerance than the wild type (wt) line to cadmium sulfide quantum dots (CdS QDs). The tolerance response was characterized at physiological, genetic and transcriptomic levels. In this work, a comparative analysis was performed on protein extracts from plantlets of the two mutants and of wt, each treated with 80 mg L−1 CdS QDs. A comparative protein analysis was performed by 2D-PAGE, and proteins were characterized by MALDI-TOF/TOF mass spectrometry. Of 250 proteins identified from all three lines, 98 showed significant changes in relative abundance between control and CdS QD-treated plantlets. The wt, atnp01, and atnp02 control-treated pairs respectively showed 61, 31, and 31 proteins with differential expression. The two mutants had a different response to treatment in terms of type and quantity of up- and downregulated proteins. This difference became more striking when compared to wt. A network analysis of the proteins differentially expressed in atnp01 and atnp02 included several of those encoded by putative genes accommodating the transposons, which were responsible for regulation of some proteins identified in this study. These included nifu-like protein 3 (Nfu3), involved in chloroplast assembly, elongator complex 3 (Elo3), involved in transcriptional elongation, magnesium-chelate subunit-2 (Chli2), involved in chlorophyll biosynthesis, and protein phosphatase 2C (PP2C) which mediates abiotic stress response.

[Display omitted] • The coffee protein profile differs before fermentation and after drying. • 2D-PAGE separation reveals significant differences in spot distribution to control. • Fermentation provides a valuable tool for modifying proteins and volatile compounds. This work aims to analyze the protein profile and volatile compounds of coffees fermented with the indigenous microbiota and with the co-inoculation of three yeasts (Saccharomyces cerevisiae , Torulaspora delbrueckii , and Candida parapsilosis). Two-dimensional gel electrophoresis (2D-PAGE), MALDI-ToF/ToF (MS/MS), and gas chromatography (GC–MS) were performed. A total of 72 "spots" were detected by 2D-PAGE. 16 spots were selected for identification by MALDI-ToF/ToF, and 12 were identified (11S protein, 13S globulin basic chain, 17.6 kDa class II heat shock protein (HSP17.6-CII), 18.0 kDa class I heat shock protein, Seed of Late Development Stage, Pru ar 1, and FAR-1 protein). 81 main volatile compounds were detected and classified into alcohols, acids, aldehydes, esters, hydrocarbons, pyrazines, furans, thiols, and pyridines/pyrrols. The difference between the identified volatile compounds and their concentrations was detected in the treatments with and without inoculation after drying. The compounds formed in green coffee during fermentation can participate in several reactions during roasting, presenting different sensory profiles and contributing to coffee quality. [ABSTRACT FROM AUTHOR]

This study was undertaken to find out the optimum physicochemical parameters of fermentation, i.e. pH, incubation temperature and incubation time for the cellulase enzyme production of Trichoderma harzianum. The extracellular protein content was estimated by the dye binding method of Bradford. Endo-glucanase (EG), exoglucanase (or Cellobiohydrolase; CBH), β-glucosidase and total cellulase activity were investigated. The molecular weight of cellulase enzymes was studied using SDS-PAGE. To identify the predominant catalytic components in optimum conditions of enzyme production, cellulases were separated by an adapted two-dimensional electrophoresis technique. Estimated optimum conditions for cellulase enzyme were found as: pH 6.5, incubation temperature 28°C and incubation time 72 h. The SDS-PAGE profiles showed several enzyme bonds such as CBHs, EGs and BGLs. The T. harzianumhad both enzyme bonds of Cel7A (CBHI) and Cel7B (EG). Finally, the results of the 2D PAGE analysis showed that the profile of protein in optimium conditions of enzyme production had several enzymes such as CBHs, EGs and the high values of cellulose activity due to synergism that occurred between the CBH and EG.

ABSTRACT Scorpions belonging to the Tityus genus are of medical interest in Brazil. Among them, Tityus stigmurus is the main scorpion responsible for stings in the Northeast region. After a sting, the scorpion venom distributes rapidly to the organs, reaching the kidneys quickly. However, there are few studies concerning the renal pathophysiology of scorpion poisoning. In this study, we evaluated the effects of T. stigmurus venom (TsV) on renal parameters in isolated rat kidneys. Wistar rats (n = 6), weighing 250-300 g, were perfused with Krebs-Henseleit solution containing 6 g/100 mL bovine serum albumin. TsV at 0.3 and 1.0 μg/mL was tested, and the effects on perfusion pressure (PP), renal vascular resistance (RVR), urinary flow (UF), glomerular filtration rate (GFR), and electrolyte excretion were analyzed. Effects were observed only at TsV concentration of 1.0 μg/mL, which increased PP (controlPP40' = 92.7 ± 1.95; TsVPP40' = 182.0 ± 4.70* mmHg, *p < 0.05), RVR (controlRVR40' = 3.28 ± 0.23 mmHg; TstRVR40' = 6.76 ± 0.45* mmHg, *p < 0.05), UF (controlUF50' = 0.16 ± 0.04; TstUF50' = 0.60 ± 0.10* mL/g/min,*p < 0.05), GFR and electrolyte excretion, with histological changes that indicate renal tubular injury. In conclusion, T. stigmurus venom induces a transient increase in PP with tubular injury, both of which lead to an augmented electrolyte excretion.

The vascular wilt of tomato caused by Fusarium oxysporum f.sp. lycopersici is an important soil borne pathogen causes severe yield loss. The molecular characterization and their interaction with its host is necessary to develop a protection strategy. 20 isolates of F. oxysporum f.sp. lycopersici (FOL) were isolated from wilt infected tomato plants across Tamil Nadu. They were subjected to cultural, morphological, molecular and virulence studies. The results revealed that all the isolates produced both micro and macro conidia with different size, number of cells. The colors of the culture and growth pattern were also varied. In addition, chlamydospores were observed terminally and intercalary. The PCR analysis with F. oxysporum species-specific primer significantly amplified an amplicon of 600 bp fragment in all the isolates. Based on the above characters and pathogenicity, isolate FOL-8 was considered as virulent and FOL-20 was considered as least virulent. Proteomics strategy was adopted to determine the virulence factors between the isolates of FOL-8 and FOL-20. The 2D analyses have showed the differential expression of 17 different proteins. Among them, three proteins were down regulated and 14 proteins were significantly up regulated in FOL-8 than FOL-20 isolate. Among the 17 proteins, 10 distinct spots were analyzed by MALDI-TOF. The functions of the analyzed proteins, suggested that they were involved in pathogenicity, symptom expression and disease development, sporulation, growth, and higher penetration rate on tomato root tissue. Overall, these experiments proves the role of proteome in pathogenicity of F. oxysporum f.sp. lycopersici in tomato and unravels the mechanism behinds the virulence of the pathogen in causing wilt disease.

Amphoteric azo dyes were used for internal control of pI values in Comparative two-dimensional Fluorescence Gel Electrophoresis (CoFGE) [1]. The 2D-gel images of separated Escherichia coli proteins as well as those of colored amphoteric dyes separated by isoelectric focussing are presented. The latter were used to correct for variation in the first electrophoretic dimension and further improve protein coordinate assignment in 2D-gel electrophoresis. Data tables are supplied to demonstrate pI-value calibration and the effect on the assignment of protein spot coordinates.

Background: Black glutinous rice contains remarkable levels of anthocyanins, which possess anti-oxidative properties and thus have health benefits. The accumulation of anthocyanins in grains of thirty black glutinous rice varieties was measured, and the results revealed that the accumulated anthocyanin content ranged from 0.262 to 2.539 mg/g. Black glutinous rice Br no. 19 was selected, and its leaf protein expression profile was compared with that of white glutinous rice RD 6 using 2D-PAGE, and the protein spots were then directly analyzed after proteolysis by LC–MS/MS. Results: The proteins from the leaves of the two rice varieties were separated using 2D-PAGE and silver stained, and the spots were analyzed using Image Master 2D Platinum version 5 software. The results showed that the protein profiles of these two rice cultivars were different, with at least six protein spots that were detected only in Br no. 19. In addition, seven protein spots accumulated at higher levels in Br no. 19 than in RD 6. Conclusion: The protein spot S1 (AP005098.4) is homologous to the Rc protein. Our results suggest that some of the proteins enriched in Br no. 19 may be involved in anthocyanin synthesis in the black glutinous rice.

This study presents data on the extraction and characterization of proteins associated with mercury in the muscle and liver tissues of jaraqui (Semaprochilodus spp.) from the Madeira River in the Brazilian Amazon. Protein fractionation was carried out by two-dimensional electrophoresis (2D-PAGE). Mercury determination in tissues, pellets, and protein spots was performed by graphite furnace atomic absorption spectrometry (GFAAS). Proteins in the spots that showed mercury were characterized by electrospray ionization tandem mass spectrometry (ESI-MS/MS). The highest mercury concentrations were found in liver tissues and pellets (426 ± 6 and 277 ± 4 μg kg−1), followed by muscle tissues and pellets (132 ± 4 and 86 ± 1 μg kg−1, respectively). Mercury quantification in the protein spots allowed us to propose stoichiometric ratios in the range of 1-4 mercury atoms per molecule of protein in the protein spots. The proteins characterized in the analysis by ESI-MS/MS were keratin, type II cytoskeletal 8, parvalbumin beta, parvalbumin-2, ubiquitin-40S ribosomal S27a, 39S ribosomal protein L36 mitochondrial, hemoglobin subunit beta, and hemoglobin subunit beta-A/B. The results suggest that proteins such as ubiquitin-40S ribosomal protein S27a, which have specific domains, possibly zinc finger, can be used as biomarkers of mercury, whereas mercury and zinc present characteristics of soft acids. [ABSTRACT FROM AUTHOR]

The vascular wilt of tomato caused by Fusarium oxysporum f.sp. lycopersici is an important soil borne pathogen causes severe yield loss. The molecular characterization and their interaction with its host is necessary to develop a protection strategy. 20 isolates of F. oxysporum f.sp. lycopersici (FOL) were isolated from wilt infected tomato plants across Tamil Nadu. They were subjected to cultural, morphological, molecular and virulence studies. The results revealed that all the isolates produced both micro and macro conidia with different size, number of cells. The colors of the culture and growth pattern were also varied. In addition, chlamydospores were observed terminally and intercalary. The PCR analysis with F. oxysporum species-specific primer significantly amplified an amplicon of 600 bp fragment in all the isolates. Based on the above characters and pathogenicity, isolate FOL-8 was considered as virulent and FOL-20 was considered as least virulent. Proteomics strategy was adopted to determine the virulence factors between the isolates of FOL-8 and FOL-20. The 2D analyses have showed the differential expression of 17 different proteins. Among them, three proteins were down regulated and 14 proteins were significantly up regulated in FOL-8 than FOL-20 isolate. Among the 17 proteins, 10 distinct spots were analyzed by MALDI-TOF. The functions of the analyzed proteins, suggested that they were involved in pathogenicity, symptom expression and disease development, sporulation, growth and higher penetration rate on tomato root tissue. Overall, these experiments proves the role of proteome in pathogenicity of F. oxysporum f.sp. lycopersici in tomato and unravels the mechanism behinds the virulence of the pathogen in causing wilt disease. [ABSTRACT FROM AUTHOR]

Mercury is a potentially toxic element that is present in the environment of the Brazilian Amazon and is responsible for adverse health effects in humans. This study sought to assess possible protein biomarkers of mercury exposure in breast milk samples from lactating women in the Madeira and Negro Rivers in the Brazilian Amazon. The mercury content of hair samples of lactating women was determined, and the proteome of breast milk samples was obtained using two-dimensional electrophoresis after protein precipitation with acetone. Mercury measurements of protein spots obtained via protein fractionation were performed by graphite furnace atomic absorption spectrometry (GFAAS), and it was observed that mercury is linked to proteins with molecular masses in the range of 14-26 kDa. The total mercury concentration was also determined by GFAAS in unprocessed milk, lyophilized milk, and protein pellets, with the purpose of determining the mercury mass balance in relation to the concentration of this element in milk and pellets. Approximately 85 to 97% of mercury present in the lyophilized milk from samples of lactating women of the Madeira River is bound in the protein fraction. From lactating women of the Negro River, approximately 49% of the total mercury is bound in the protein fraction, and a difference of 51% is bound in the lipid fraction. [ABSTRACT FROM AUTHOR]

Proteomic methods allow detecting breed-specific differences at protein level. The objective of the present study was to compare muscle proteome profiles of Hungarian Merino and Tsigai sheep breeds by two dimensional gel electrophoresis and mass spectrometry based protein identification. Approximately 327 protein spots were detected in musculus longissimus dorsi and 14 protein spots, identified as 12 proteins, showed significant differences in their intensity (P < 0.05) between breeds. Seven and five proteins had higher expression in Merino and Tsigai skeletal muscle, respectively. Identified proteins were classified as structural, carbohydrate metabolism-related and miscellaneous ones. Six genes, which represent the three protein groups by function, were analysed by quantitative real-time PCR. All of the identified structural proteins have shown higher intensity in Merino, while the expression of identified glycolytic enzymes and myoglobin were higher in Tsigai lambs. One of the structural and one of the carbohydrate metabolism-related protein showed differences in expression at mRNA level by quantitative real-time PCR, as well. To the best of our knowledge, it was the first differential proteome analysis of sheep muscle and these results contribute to a better understanding of molecular differences between breeds. [ABSTRACT FROM AUTHOR]