Subcellular proteomic characterization of the high-temperature stress response of the cyanobacterium Spirulina platensis

The present study examined the changes in protein expression in Spirulina platensis upon exposure to high temperature, with the changes in expression analyzed at the subcellular level. In addition, the transcriptional expression level of some differentially expressed proteins, the expression pattern clustering, and the protein-protein interaction network were analyzed. The results obtained from differential expression analysis revealed up-regulation of proteins involved in two-component response systems, DNA damage and repair systems, molecular chaperones, known stress-related proteins, and proteins involved in other biological processes, such as capsule formation and unsaturated fatty acid biosynthesis. The clustering of all differentially expressed proteins in the three cellular compartments showed: (i) the majority of the proteins in all fractions were sustained tolerance proteins, suggesting the roles of these proteins in the tolerance to high temperature stress, (ii) the level of resistance proteins in the photosynthetic membrane was 2-fold higher than the level in two other fractions, correlating with the rapid inactivation of the photosynthetic system in response to high temperature. Subcellular communication among the three cellular compartments via protein-protein interactions was clearly shown by the PPI network analysis. Furthermore, this analysis also showed a connection between temperature stress and nitrogen and ammonia assimilation

Comparative analysis of the Spirulina platensis subcellular proteome in response to low- and high-temperature stresses: uncovering cross-talk of signaling components

The present study focused on comparative proteome analyses of low- and high-temperature stresses and potential protein-protein interaction networks, constructed by using a bioinformatics approach, in response to both stress conditions

Desaturase specificity is controlled by the physicochemical properties of a single amino acid residue in the substrate binding tunnel

Membrane fatty acyl desaturases (mFAD) are ubiquitous enzymes in eukaryotes. They introduce double bonds into fatty acids (FAs), producing structurally diverse unsaturated FAs which serve as membrane lipid components or precursors of signaling molecules. The mechanisms controlling enzymatic specificity and selectivity of desaturation are, however, poorly understood. We found that the physicochemical properties, particularly side chain volume, of a single amino acid (aa) residue in insect mFADs (Lepidoptera: Bombyx mori and Manduca sexta) control the desaturation products. Molecular dynamics simulations of systems comprising wild-type or mutant mFADs with fatty acyl-CoA substrates revealed that the single aa substitution likely directs the outcome of the desaturation reaction by modulating the distance between substrate fatty acyl carbon atoms and active center metal ions. These findings, as well as our methodology combining mFAD mutational screening with molecular dynamics simulations, will facilitate prediction of desaturation products and facilitate engineering of mFADs for biotechnological applications

Microalgae biomass as an alternative ingredient in cookies: sensory, physical and chemical properties, antioxidant activity and in vitro digestibility

Microalgae can be regarded as an alternative and promising food ingredient due to their nutritional composition, richness in bioactive compounds, and because they are considered a sustainable protein source for the future. The aim of this work was to evaluate microalgae (Arthrospira platensis F & M-C256, Chlorella vulgaris Allma, Tetraselmis suecica F & M-M33 and Phaeodactylum tricornutum F & M-M40) as innovative ingredients to enhance functional properties of cookies. Two biomass levels were tested and compared to control: 2% (w/w) and 6% (w/ w), to provide high levels of algae-bioactives. The cookies sensory and physical properties were evaluated during eight weeks showing high color and texture stability. Cookies prepared with A. platensis and C. vulgaris presented significantly (p < 0.05) higher protein content compared to the control, and by sensory analysis A. platensis cookies were preferred. Besides, A. platensis also provided a structuring effect in terms of cookies texture. All microalgae-based cookies showed significantly higher (p < 0.05) total phenolic content and in vitro antioxidant capacity compared to the control. No significant difference (p < 0.05) in in vitro digestibility between microalgae cookies and the control was foundinfo:eu-repo/semantics/publishedVersio

Proteome-wide analysis and diel proteomic profiling in the cyanobacterium Arthrospira platensis PCC 8005

The filamentous cyanobacteriumArthrospira platensishas a long history of use as a food supply and it has been used by the European Space Agency in the MELiSSA project, an artificial microecosystem which supports life during long-term manned space missions. This study assesses progress in the field of cyanobacterial shotgun proteomics and light/dark diurnal cycles by focusing onArthrospira platensis. Several fractionation workflows including gel-free and gel-based protein/peptide fractionation procedures were used and combined with LC-MS/MS analysis, enabling the overall identification of 1306 proteins, which represents 21% coverage of the theoretical proteome. A total of 30 proteins were found to be significantly differentially regulated under light/dark growth transition. Interestingly, most of the proteins showing differential abundance were related to photosynthesis, the Calvin cycle and translation processes. A novel aspect and major achievement of this work is the successful improvement of the cyanobacterial proteome coverage using a 3D LC-MS/MS approach, based on an immobilized metal affinity chromatography, a suitable tool that enabled us to eliminate the most abundant protein, the allophycocyanin. We also demonstrated that cell growth follows a light/dark cycle inA. platensis. This preliminary proteomic study has highlighted new characteristics of theArthrospira platensisproteome in terms of diurnal regulation

Assembly of Pseudomonas putida Aspartate Transcarbamoylase and Possible Roles of the PyrC' Polypeptide in the Folding of the Dodecameric Enzyme

Aspartate transcarbamoylase (ATCase) of Pseudomonas putida consists of two different polypeptides, PyrB and PyrC' (Schurr et al, 1995). The role of the PyrC' and the assembly of PyrB and PyrC' have been studied. The ATCase made in vitro of P.putida PyrB with P.putida PyrC', and of E.coli PyrB with P.putida PyrC ' were generated under two different conditions, denaturation and renaturation, and untreated. It was found that PyrC' plays a role in the enzymatic regulation by ATP, CTP and UTP. In addition to playing a role in substrate binding, the PyrB polypeptide is also involved in effector binding (Kumar et al., manuscript in preparation). The most energetically preferred form of the P.putida WT is a dodecamer with a molecular mass of 480 kDa. The ratio between the PyrB and the PyrC' is 1:1. In studies of nucleotide binding, it was discovered that the P.putida PyrB was phosphorylated by a protein kinase in the cell extract. In the presence of 20 mM EDTA, this phosphorylation was inhibited and the inhibition could be overcome by the addition of divalent cations such as Zn2+ and Mg2+. This result suggested that the phosphorylation reaction required divalent cations. In the CAD complex of eukaryotes, phosphorylations of the CPSase and the linker region between ATCase and DHOase did not occur in the presence of UTP and it was hypothesized (Carrey, 1993) that UTP and phosphorylation(s) regulated the conformational change in the enzyme complex. Therefore, the same idea was approached with P.putida ATCase, where it was found that 1.0 mM UTP inhibited the phosphorylation of PyrB by more than 50%. These results suggested that the regulation of the conformational change of the P.putida ATCase might be similar to that of CAD. Furthermore, peptide mapping for phosphorylation sites was performed on P.putida ATCase WT, WT --11 amino acids and WT --34 amino acids from the N-terminus of the PyrB polypeptide. The results showed that the phosphorylation sites were located on the fragment that contained amino acid number-35 to amino acid number-112 from the N-terminus of the PyrB polypeptide

Ensemble-AMPPred: Robust AMP Prediction and Recognition Using the Ensemble Learning Method with a New Hybrid Feature for Differentiating AMPs

Antimicrobial peptides (AMPs) are natural peptides possessing antimicrobial activities. These peptides are important components of the innate immune system. They are found in various organisms. AMP screening and identification by experimental techniques are laborious and time-consuming tasks. Alternatively, computational methods based on machine learning have been developed to screen potential AMP candidates prior to experimental verification. Although various AMP prediction programs are available, there is still a need for improvement to reduce false positives (FPs) and to increase the predictive accuracy. In this work, several well-known single and ensemble machine learning approaches have been explored and evaluated based on balanced training datasets and two large testing datasets. We have demonstrated that the developed program with various predictive models has high performance in differentiating between AMPs and non-AMPs. Thus, we describe the development of a program for the prediction and recognition of AMPs using MaxProbVote, which is an ensemble model. Moreover, to increase prediction efficiency, the ensemble model was integrated with a new hybrid feature based on logistic regression. The ensemble model integrated with the hybrid feature can effectively increase the prediction sensitivity of the developed program called Ensemble-AMPPred, resulting in overall improvements in terms of both sensitivity and specificity compared to those of currently available programs

SpirPep: an in silico digestion-based platform to assist bioactive peptides discovery from a genome-wide database

Abstract Background Bioactive peptides, including biological sources-derived peptides with different biological activities, are protein fragments that influence the functions or conditions of organisms, in particular humans and animals. Conventional methods of identifying bioactive peptides are time-consuming and costly. To quicken the processes, several bioinformatics tools are recently used to facilitate screening of the potential peptides prior their activity assessment in vitro and/or in vivo. In this study, we developed an efficient computational method, SpirPep, which offers many advantages over the currently available tools. Results The SpirPep web application tool is a one-stop analysis and visualization facility to assist bioactive peptide discovery. The tool is equipped with 15 customized enzymes and 1–3 miscleavage options, which allows in silico digestion of protein sequences encoded by protein-coding genes from single, multiple, or genome-wide scaling, and then directly classifies the peptides by bioactivity using an in-house database that contains bioactive peptides collected from 13 public databases. With this tool, the resulting peptides are categorized by each selected enzyme, and shown in a tabular format where the peptide sequences can be tracked back to their original proteins. The developed tool and webpages are coded in PHP and HTML with CSS/JavaScript. Moreover, the tool allows protein-peptide alignment visualization by Generic Genome Browser (GBrowse) to display the region and details of the proteins and peptides within each parameter, while considering digestion design for the desirable bioactivity. SpirPep is efficient; it takes less than 20 min to digest 3000 proteins (751,860 amino acids) with 15 enzymes and three miscleavages for each enzyme, and only a few seconds for single enzyme digestion. Obviously, the tool identified more bioactive peptides than that of the benchmarked tool; an example of validated pentapeptide (FLPIL) from LC-MS/MS was demonstrated. The web and database server are available at http://spirpepapp.sbi.kmutt.ac.th. Conclusion SpirPep, a web-based bioactive peptide discovery application, is an in silico-based tool with an overview of the results. The platform is a one-stop analysis and visualization facility; and offers advantages over the currently available tools. This tool may be useful for further bioactivity analysis and the quantitative discovery of desirable peptides