scholarly journals Simultaneous Production of Multiple Antimicrobial Compounds by Bacillus velezensis ML122-2 Isolated From Assam Tea Leaf [Camellia sinensis var. assamica (J.W.Mast.) Kitam.]

2021 ◽  
Vol 12 ◽  
Author(s):  
Patthanasak Rungsirivanich ◽  
Elvina Parlindungan ◽  
Paula M. O’Connor ◽  
Des Field ◽  
Jennifer Mahony ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Camellia Sinensis ◽  
Gene Clusters ◽  
Antimicrobial Activities ◽  
Transcriptional Analysis ◽  
Mass Spectrometry Analysis ◽  
Bacillus Velezensis ◽  
Spectrometry Analysis ◽  
Wide Range ◽  
Assam Tea

Bacillus velezensis ML122-2 is an antimicrobial-producing strain isolated from the leaf of Assam tea or Miang [Camellia sinensis var. assamica (J.W.Mast.) Kitam.]. The cell-free supernatant (CFS) of strain ML122-2 exhibits a broad-spectrum antimicrobial activity against various Gram-positive and Gram-negative bacteria as well as the mold Penicillium expansum. The genome of B. velezensis ML122-2 was sequenced and in silico analysis identified three potential bacteriocin-associated gene clusters, that is, those involved in the production of mersacidin, amylocyclicin, and LCI. Furthermore, six gene clusters exhibiting homology (75–100% DNA sequence identity) to those associated with the secondary metabolites bacilysin, bacillibactin, surfactin, macrolactin H, bacillaene, and plipastatin were identified. Individual antimicrobial activities produced by B. velezensis ML122-2 were purified and characterized by Matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometry analysis, revealing three antimicrobial peptides with molecular masses corresponding to surfactin, plipastatin, and amylocyclicin. Transcriptional analysis of specific genes associated with mersacidin (mrsA), amylocyclicin (acnA), plipastatin (ppsA), and surfactin (srfAA) production by B. velezensis ML122-2 showed that the first was not transcribed under the conditions tested, while the latter three were consistent with the presence of the associated peptides as determined by mass spectrometry analysis. These findings demonstrate that B. velezensis ML122-2 has the genetic capacity to produce a wide range of antimicrobial activities that may support a specific community structure and highlight the biotechnological properties of Assam tea.

scholarly journals Novel exosome biomarker candidates for Alzheimer´s disease unraveled through Mass Spectrometry analysis

2021 ◽  
Author(s):  
Tânia Soares Martins ◽  
Rui Marçalo ◽  
Cristóvão B. da Cruz e Silva ◽  
Dário Trindade ◽  
José Catita ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Multivariate Analyses ◽  
Cost Effective ◽  
Accurate Diagnosis ◽  
Functional Enrichment ◽  
Mass Spectrometry Analysis ◽  
Spectrometry Analysis ◽  
Wide Range ◽  
Go Terms ◽  
Peripheral Biomarker

Abstract Exosomes are small extracellular vesicles (EVs) present in human biofluids that can transport specific disease-associated molecules. Consequently blood-derived exosomes have emerged as important peripheral biomarker sources for a wide range of diseases, among them Alzheimer’s disease (AD). Although there is no effective cure for AD, an accurate diagnosis, relying on easily accessible peripheral biofluids, is still necessary to discriminate this disease from other dementias, test potential therapies and even monitor rate of disease progression. The ultimate goal is to produce a cost-effective and widely available alternative, which can also be employed as a first clinical screen. In this study, EVs with exosome-like characteristics were isolated from serum of Controls and AD cases through precipitation- and column-based methods, followed by mass spectrometry analysis. The resulting proteomes were characterized by Gene Ontology (GO) functional enrichment and multivariate analyses. Although GO terms were similar for exosomes proteomes of Controls and ADs, using both methodologies, a clear segregation of disease cases was obtained when using the precipitation-based method. Nine significantly different abundant proteins were identified between Controls and AD cases, representing putative biomarker candidate targets. Among them AACT and C4BPα, two Aβ-binding proteins, whose exosome levels were further validated in individuals from independent cohorts using antibody-based approaches. The findings discussed represent an important contribution to the identification of novel exosomal biomarker candidates useful as potential blood-based tools for AD diagnosis.

scholarly journals Novel exosome biomarker candidates for Alzheimer´s disease unraveled through Mass Spectrometry analysis

2021 ◽  
Author(s):  
Tânia Soares Martins ◽  
Rui Marçalo ◽  
Cristóvão B. da Cruz e Silva ◽  
Dário Trindade ◽  
José Catita ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Multivariate Analysis ◽  
Cost Effective ◽  
Functional Enrichment ◽  
Mass Spectrometry Analysis ◽  
Precipitation Method ◽  
Molecular Diagnostic ◽  
Diagnostic Tools ◽  
Spectrometry Analysis ◽  
Wide Range

Abstract Background Exosomes are small extracellular vesicles (EVs) present in human biofluids that can carry disease-specific molecules. Blood-derived exosomes emerged as interesting peripheral sources of biomarkers for a wide range of diseases, and their potential is also being addressed in Alzheimer’s disease (AD) context. There is no effective cure or blood-based molecular diagnostic tools for a first clinical AD screening, which motivates research in this area to identify putative valuable blood-derived disease biomarkers. The ultimate goal is to produce a cost-effective and widely available alternative to the current molecular AD diagnosis that monitors a biomarker triplet in the CSF. Methods In the study here presented, EVs with exosome-like characteristics were isolated from the serum of Controls and AD cases through two distinct methods, precipitation- and column-based methods, followed by mass spectrometry analysis. The resulting proteomes from Controls and AD cases were characterized by Gene Ontology (GO) functional enrichment and multivariate analysis. Putative candidate targets identified were validated in distinct cohorts using antibody-based approaches. Results Both methodologies isolated particles with the expected morphology and size range. Although GO terms were similar for Controls and AD cases exosomes in both methodologies, the multivariate analysis revealed a clear segregation between Controls and AD cases obtained by the precipitation method. Nine significantly different abundant proteins were identified between Controls and AD cases and exosome levels of AACT and C4BPα, two Aβ-binding proteins, were validated in individuals from independent cohorts. Conclusions Serum-derived exosome proteomes were unraveled for the first time in the context of AD through two distinct isolation methodologies, holding disease discriminatory potential. The work carried out gives an important contribution to the identification of novel exosomal biomarker candidates potentially useful as blood-based tools for AD diagnosis.

scholarly journals From simple and specific zymographic detections to the annotation of a fungus Daldinia caldariorum D263 that encodes a wide range of highly bioactive cellulolytic enzymes

2021 ◽  
Vol 14 (1) ◽  
Author(s):  
Meng-Chun Lin ◽  
Hsion-Wen Kuo ◽  
Mu-Rong Kao ◽  
Wen-Dar Lin ◽  
Chen-Wei Li ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Endophytic Fungus ◽  
Agricultural Waste ◽  
Fungal Species ◽  
Mass Spectrometry Analysis ◽  
Cellulolytic Enzymes ◽  
Cellulolytic Enzyme ◽  
Specific Method ◽  
Spectrometry Analysis ◽  
Wide Range

Abstract Background Lignocellulolytic enzymes are essential for agricultural waste disposal and production of renewable bioenergy. Many commercialized cellulase mixtures have been developed, mostly from saprophytic or endophytic fungal species. The cost of complete cellulose digestion is considerable because a wide range of cellulolytic enzymes is needed. However, most fungi can only produce limited range of highly bioactive cellulolytic enzymes. We aimed to investigate a simple yet specific method for discovering unique enzymes so that fungal species producing a diverse group of cellulolytic enzymes can be identified. Results The culture medium of an endophytic fungus, Daldinia caldariorum D263, contained a complete set of cellulolytic enzymes capable of effectively digesting cellulose residues into glucose. By taking advantage of the unique product inhibition property of β-glucosidases, we have established an improved zymography method that can easily distinguish β-glucosidase and exoglucanase activity. Our zymography method revealed that D263 can secrete a wide range of highly bioactive cellulases. Analyzing the assembled genome of D263, we found over 100 potential genes for cellulolytic enzymes that are distinct from those of the commercially used fungal species Trichoderma reesei and Aspergillus niger. We further identified several of these cellulolytic enzymes by mass spectrometry. Conclusions The genome of Daldinia caldariorum D263 has been sequenced and annotated taking advantage of a simple yet specific zymography method followed by mass spectrometry analysis, and it appears to encode and secrete a wide range of bioactive cellulolytic enzymes. The genome and cellulolytic enzyme secretion of this unique endophytic fungus should be of value for identifying active cellulolytic enzymes that can facilitate conversion of agricultural wastes to fermentable sugars for the industrial production of biofuels.

scholarly journals Effect of Ocular Hypertension on D-β-Aspartic Acid-Containing Proteins in the Retinas of Rats

2019 ◽  
Vol 2019 ◽  
pp. 1-8 ◽  
Author(s):  
Takashi Kanamoto ◽  
Takashi Tachibana ◽  
Yasushi Kitaoka ◽  
Toshio Hisatomi ◽  
Yasuhiro Ikeda ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Ocular Hypertension ◽  
Aspartic Acid ◽  
Atp Synthase ◽  
Wistar Rats ◽  
Protein Band ◽  
Mass Spectrometry Analysis ◽  
Liquid Chromatography Mass Spectrometry ◽  
Spectrometry Analysis ◽  
Sds Page

Purpose. To investigate the effect of ocular hypertension-induced isomerization of aspartic acid in retinal proteins. Methods. Adult Wistar rats with ocular hypertension were used as an experimental model. D-β-aspartic acid-containing proteins were isolated by SDS-PAGE and western blot with an anti-D-β-aspartic acid antibody and identified by liquid chromatography-mass spectrometry analysis. The concentration of ATP was measured by ELISA. Results. D-β-aspartic acid was expressed in a protein band at around 44.5 kDa at much higher quantities in the retinas of rats with ocular hypertension than in those of normotensive rats. The 44.5 kDa protein band was mainly composed of α-enolase, S-arrestin, and ATP synthase subunits α and β, in both the ocular hypertensive and normotensive retinas. Moreover, increasing intraocular pressure was correlated with increasing ATP concentrations in the retinas of rats. Conclusion. Ocular hypertension affected the expression of proteins containing D-β-aspartic acid, including ATP synthase subunits, and up-regulation of ATP in the retinas of rats.

scholarly journals N-Terminomics Strategies for Protease Substrates Profiling

Molecules ◽  
2021 ◽  
Vol 26 (15) ◽  
pp. 4699
Author(s):  
Mubashir Mintoo ◽  
Amritangshu Chakravarty ◽  
Ronak Tilvawala
Keyword(s):  
Mass Spectrometry ◽  
Protease Activity ◽  
Viral Infections ◽  
Mass Spectrometry Analysis ◽  
Post Translational Modification ◽  
Spectrometry Analysis ◽  
Physiological Conditions ◽  
Inflammatory Conditions ◽  
Biological Mixtures

Proteases play a central role in various biochemical pathways catalyzing and regulating key biological events. Proteases catalyze an irreversible post-translational modification called proteolysis by hydrolyzing peptide bonds in proteins. Given the destructive potential of proteolysis, protease activity is tightly regulated. Dysregulation of protease activity has been reported in numerous disease conditions, including cancers, neurodegenerative diseases, inflammatory conditions, cardiovascular diseases, and viral infections. The proteolytic profile of a cell, tissue, or organ is governed by protease activation, activity, and substrate specificity. Thus, identifying protease substrates and proteolytic events under physiological conditions can provide crucial information about how the change in protease regulation can alter the cellular proteolytic landscape. In recent years, mass spectrometry-based techniques called N-terminomics have become instrumental in identifying protease substrates from complex biological mixtures. N-terminomics employs the labeling and enrichment of native and neo-N-termini peptides, generated upon proteolysis followed by mass spectrometry analysis allowing protease substrate profiling directly from biological samples. In this review, we provide a brief overview of N-terminomics techniques, focusing on their strengths, weaknesses, limitations, and providing specific examples where they were successfully employed to identify protease substrates in vivo and under physiological conditions. In addition, we explore the current trends in the protease field and the potential for future developments.

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