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Peptide profile of the storage proteins of chickpea, wheat, soybean analyzed with LC-MS/MS

ABSTRACT: Aiming to reduce food spoilage, the present study developed novel highly active food-grade preservatives affecting a wide range of bacteria. For this purpose, storage proteins were extracted from food plants. After enzymatic hydrolysis by the digestive protease chymotrypsin, the peptide profiles were analyzed by ultrahigh-performance micro-liquid chromatography–triple quadrupole time-of-flight tandem mass spectrometry. Virtual screening identified 21 potential antimicrobial peptides in chickpea legumin. Among those, the peptides Leg1 (RIKTVTSFDLPALRFLKL) and Leg2 (RIKTVTSFDLPALRWLKL) exhibited antimicrobial activity against 16 different bacteria, including pathogens, spoilage-causing bacteria and two antibiotic-resistant strains. Minimum inhibitory concentrations (MIC) down to 15.6 µM indicated 10–1,000-fold higher activity of the novel antimicrobial peptides compared to conventional food preservatives. Moreover, Leg1 and Leg2 showed bactericidal activity in bacterial suspension and during the storage of raw pork meat.

INSTRUMENT(S): TripleTOF 6600

ORGANISM(S): Cicer Arietinum (chickpea) (garbanzo) Triticum Aestivum (wheat) Glycine Max

SUBMITTER: Marie-Louise Heymich

LAB HEAD: Monika Pischetsrieder

PROVIDER: PXD019257 | Pride | 2021-01-11

REPOSITORIES: Pride

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Project description:Background. Silkworm pupae (SWP) is the main by-product of the sericulture industry with an interesting nutritional profile, especially in terms of proteins. In consideration of its possible use as a food or food ingredient in Western countries, a comparative proteomic experiment has been performed to investigate the differences of the protein profile of male and female SWP reared on mulberry leaves or on an artificial diet. Methods. The nutritional profile of lyophilized SWP in terms of dry matter and ash was evaluated according to the AOAC procedures, the total nitrogen content was determined by a nitrogen analyzer and the SWP gross energy value was measured using an adiabatic calorimetric bomb. The comparative proteomic analysis was performed on male and female SWP reared on mulberry leaves or on the artificial diet. Proteins were separated by Bidimensional Electrophoresis (2DE) and, after a multivariate statistical analysis, the differentially expressed proteins were identified by LC-MS/MS. Results. The comparative proteomic approach highlighted 47 SWP proteins differentially expressed comparing diet and gender. PCA analysis showed that 7 proteins were more effective in discriminating the sex and 5 were more effective in discriminating the diet type. In spite of the above mentioned differences in the SWP protein profile, no strong alteration of the pupa physiological traits have been demonstrated, suggesting a general SWP flexibility to adapt to a well-balanced artificial diet. Differences in lipid transport and metabolism were found among the experimental groups, that might have a relevant effect on the timing, on hormone secretion and, in turn, on insect voltinism. This aspect may also affect silk production, as univoltine strains are the most productive. Although this is a preliminary study, the proteomic data may offer a contribution in understanding also the influence of gender and farming strategy on the allergen profile of B. mori, when used as food or as a food ingredient. Since female silkworm pupae reared on mulberry leaves seemed to contain lower levels of known allergens than those reared in the other experimental conditions, we speculated that these findings may help when farming B. mori for food production purposes. However, these results need to be supported by further characterization of the allergenic potential of B. mori.

Project description:Emerging antibiotic resistance among clinically relevant bacteria, paired with their ability to form biofilms on medical and technical devices, represents a serious problem in terms of effective and long-term decontamination in health care environments and gives rise to an urgent need for new antimicrobial materials. Here we present the first study of the impact of AGXXÂ®, a novel broad-spectrum antimicrobial surface coating consisting of micro galvanic elements formed by silver and ruthenium, on the transcriptome of the nosocomial pathogen Enterococcus faecalis. E. faecalis was subjected to metal stress by growing it for different periods of time in the presence of AGXXÂ® or silver-coated steel meshes. Subsequently, total RNA was isolated and next-generation RNA sequencing was performed to analyze variations in gene expression levels in the presence of the antimicrobial materials with focus on known stress genes. Exposure to AGXXÂ® had a large impact on the transcriptome of E. faecalis. After 24 minutes almost 1/5 of the E. faecalis genome displayed differential expression. At each time-point the cop operon was strongly up-regulated, providing indirect evidence for the presence of free Ag+-ions. Moreover, exposure to AGXXÂ® induced a broad general stress response in E. faecalis. Genes coding for the chaperones GroEL and GroES as well as the Clp proteases ClpE and ClpB were among the top up-regulated heat shock genes. Furthermore, differential expression of genes coding for thioredoxin, superoxide dismutase and glutathione synthetase indicates a high level of oxidative stress. We postulate a mechanism of action where the combination of Ag+-ions and reactive oxygen species generated by AGXXÂ® results in a synergistic antimicrobial effect, which is superior to that of conventional silver coatings. Gene expression analysis of Enterococcus faecalis 12030 either subjected to metal stress by exposure to an antimicrobial AGXXÂ®- or Ag-coated V2A steel mesh or exposed to an uncoated V2A steel mesh or left untreated performing RNA Sequencing with an Ion ProtonTM Sequencer and subsequent data analysis with a T-REx RNA-Sequencing expression analysis pipeline.

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Peptide profile of the storage proteins of chickpea, wheat, soybean analyzed with LC-MS/MS

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