Proteomic investigation reveals the role of bacterial laccase from Bacillus pumilus in oxidative stress defense.

The wide distribution of laccases in nature makes them involved in different biological processes. However, little information is known about how laccase participates in the defense machinery of bacteria against oxidative stress. The present study aimed to elucidate the oxidative stress response mechanism of Bacillus pumilus ZB1 and the functional role of bacterial laccase in stress defense. The oxidative stress caused by methyl methanesulfonate (MMS) significantly induced laccase activity and its transcript level. The morphological analysis revealed that the defense of B. pumilus ZB1 against oxidative stress was activated. Based on the proteomic study, 114 differentially expressed proteins (DEPs) were up-regulated and 79 DEPs were down-regulated. In COG analysis, 66.40% DEPs were classified into the category "Metabolism". We confirmed that laccase was up-regulated in response to MMS stress and its functional annotation was related to "Secondary metabolites biosynthesis, transport and catabolism". Based on protein-protein interaction prediction, two up-regulated DEPs (YcnJ and GabP) showed interaction with laccase and contributed to the formation of laccase stability and adaptability. The overexpressed laccase might improve the antioxidative property of B. pumilus ZB1. These findings provide an insight and the guidelines for better exploitation of bioremediation using bacterial laccase. SIGNIFICANCE: Bacillus pumilus is a gram-positive bacterium that has the potential for many applications, such as bioremediation. The expression of bacterial laccase is significantly influenced by oxidative stress, while the underlying mechanism of laccase overexpression in bacteria has not been fully studied. Elucidation of the biological process may benefit the bioremediation using bacteria in the future. In this study, the differentially expressed proteins were analyzed using a TMT-labeling proteomic approach when B. pumilus was treated with methyl methanesulfonate (MMS). Reactive oxygen species induced by MMS activated the secondary metabolites biosynthesis, transport, and catabolism in B. pumilus, including laccase overexpression. Moreover, the simultaneously up-regulated YcnJ and GabP may benefit the synthesis and the stability of laccase, then improve the antioxidative property of B. pumilus against environmental stress. Our findings advance the understanding of the adaptive mechanism of B. pumilus to environmental conditions.

Enhanced denitrification of biodegradable polymers using Bacillus pumilus in aerobic denitrification bioreactors: Performance and mechanism.

Nitrate accumulation is an important issue that affects animal health and causes eutrophication. This study combined biodegradable polymers with degrading bacteria to lead to high denitrification efficiency. The results showed polycaprolactone had the highest degradation and carbon release rate (0.214 mg/g∙d) and nitrogen removal was greatest when the Bacillus pumilus and Halomonas venusta ratio was 1:2. When the hydraulic retention time was extended to 12 h, the nitrate removal rate for H. venusta with B. pumilus and polycaprolactone increased by 48 %. Furthermore, the group with B. pumilus contained more Proteobacteria (77.34 %) and denitrifying functional enzymes than the group without B. pumilus. These findings indicated B.pumilus can enhance the degradation of biodegradable polymers especially polycaprolactone to improve the denitrification of the aerobic denitrification bacteria H.venusta when treating maricultural wastewater.

Characterization of pumilacidin, a lipopeptide biosurfactant produced from Bacillus pumilus NITDID1 and its prospect in bioremediation of hazardous pollutants.

Highly hydrophobic compounds like petroleum and their byproducts, once released into the environment, can persist indefinitely by virtue of their ability to resist microbial degradation, ultimately paving the path to severe environmental pollution. Likewise, the accumulation of toxic heavy metals like lead, cadmium, chromium, etc., in the surroundings poses an alarming threat to various living organisms. To remediate the matter in question, the applicability of a biosurfactant produced from the mangrove bacterium Bacillus pumilus NITDID1 (Accession No. KY678446.1) is reported here. The structural characterization of the produced biosurfactant revealed it to be a lipopeptide and has been identified as pumilacidin through FTIR, NMR, and MALDI-TOF MS. The critical micelle concentration of pumilacidin was 120 mg/L, and it showed a wide range of stability in surface tension reduction experiments under various environmental conditions and exhibited a high emulsification index of as much as 90%. In a simulated setup of engine oil-contaminated sand, considerable oil recovery (39.78%) by this biosurfactant was observed, and upon being added to a microbial consortium, there was an appreciable enhancement in the degradation of the used engine oil. As far as the heavy metal removal potential of biosurfactant is concerned, as much as 100% and 82% removal was observed for lead and cadmium, respectively. Thus, in a nutshell, the pumilacidin produced from Bacillus pumilus NITDID1 holds promise for multifaceted applications in the field of environmental remediation.

Prediction and validation of enzymatic degradation of aflatoxin M1: Genomics and proteomics analysis of Bacillus pumilus E-1-1-1 enzymes.

Aflatoxins are a class of highly toxic mycotoxins. Aflatoxin M1 (AFM1) is hydroxylated metabolite of aflatoxin B1, having comparable toxicity, which is more commonly found in milk. In this study, the whole genome sequencing of Bacillus pumilus E-1-1-1 isolated from feces of 38 kinds of animals, having aflatoxin M1 degradation ability was conducted. Bacterial genome sequencing indicated that a total of 3445 sequences were finally annotated on 23 different cluster of orthologous groups (COG) categories. Then, the potential AFM1 degradation proteins were verified by proteomics; the properties of these proteins were further explored, including protein molecular weight, hydrophobicity, secondary structure prediction, and three-dimensional structures. Bacterial genome sequencing combined with proteomics showed that eight genes were the most capable of degrading AFM1 including three catalases, one superoxide dismutase, and four peroxidases to clone. These eight genes with AFM1 degrading capacity were successfully expressed. These results indicated that AFM1 can be degraded by Bacillus pumilus E-1-1-1 protein and the most degrading proteins were oxidoreductases.

Dynamics of bacterial community, metabolites profile and physicochemical characteristics during solid-state fermentation of soybean meal and corn mixed substrates inoculated with Bacillus pumilus and Limosilactobacillus fermentum.

BACKGROUND: Solid-state fermentation (SSF) is a general approach for preparing food and feed, which not only improves nutrition but also provides prebiotics and metabolites. Although many studies have been conducted on the effects of fermentation on feed substrate, the dynamics of microbiota and metabolites in SSF remain unclear. Here, high-throughput sequencing combined with gas chromatography-quadrupole time-of-flight mass spectrometry was used to evaluate the dynamic changes of solid fermented soybean meal and corn mixed matrix inoculated with Bacillus pumilus and Limosilactobacillus fermentum. RESULTS: Generally, inoculated bacteria rapidly proliferated, accompanied by the degradation of macromolecular proteins and an increase in the content of small peptides, trichloroacetic acid-soluble protein, free amino acids and organic acids. Bacillus, Lactobacillus and Enterococcus dominated the whole fermentation process. 389 non-volatile metabolites and 182 volatile metabolites were identified, including amino acids, organic acids, ketones, aldehydes, furans and pyrazine. Typical non-volatile metabolites such as lactic acid, 4-aminobutanoic acid, l-glutamic acid, d-arabinose and volatile metabolites such as 4-ethyl-2-methoxyphenol, 4-penten-2-ol, 2-pentanone, 2-ethylfuran, 2-methylhexanoic acid and butanoic acid-ethyl ester were significantly increased in two-stage solid fermentation. However, some adverse metabolites were also produced, such as oxalic acid, acetic acid, tyramine and n-butylamine, which may affect the quality of fermented feed. Sixteen genera were significantly correlated with differential non-volatile metabolites, while 11 genera were significantly correlated with differential volatile metabolites. CONCLUSION: These results characterized the dynamic changes in the process of two-stage solid-state fermentation with Bacillus pumilus and Limosilactobacillus fermentum and provided a potential reference for additional intervention on improving the effectiveness and efficiency of solid-state fermentation of feed in the future. © 2023 Society of Chemical Industry.

Transcriptome analysis reveals the regulatory effects of Bacillus amyloliquefaciens and Bacillus pumilus on immune and digestive related genes in the spleen of weanling black goats.

The aim of the present study was to evaluate the effects of Bacillus amyloliquefaciens fsznc-06 and Bacillus pumilus fsznc-09 on the expressions of spleen genes in weanling Jintang black goats. Bacillus amyloliquefaciens fsznc-06 (BA-treated group) and Bacillus pumilus fsznc-09 (BP-treated group) were directly fed to goats, and the spleens were harvested for transcriptome analysis. The KEGG pathway analysis showed that the differentially expressed genes (DEGs) in BA-treated vs CON group were mainly involved in digestive system and immune system, while those in BP-treated vs CON group were mainly involved in immune system, and those in BA-treated vs BP-treated group were mainly involved in digestive system. In conclusion, Bacillus amyloliquefaciens fsznc-06 might promote the expressions of genes related to immune system and digestive system, reduce the expressions of disease genes related to digestive system and might promote mutual accommodation of some immune genes in weanling black goat. Bacillus pumilus fsznc-09 might promote the expressions of genes related to immune system and mutual accommodation of some immune genes in weanling black goat. Bacillus amyloliquefaciens fsznc-06 has advantages over Bacillus pumilus fsznc-09 in promoting the expressions of genes related to digestive system and mutual accommodation of some immune genes.

Modulating the production of xylanase by Bacillus pumilus BS131 through optimization using waste fiber sludge / Modulando a produção de xilanase por Bacillus pumilus BS131 por meio da otimização usando lodo de fibra residual

Abstract In recent days, cheapest alternative carbon source for fermentation purpose is desirable to minimize production cost. Xylanases have become attractive enzymes as their potential in bio-bleaching of pulp and paper industry. The objective of the present study was to identify the potential ability on the xylanase production by locally isolated Bacillus pumilus BS131 by using waste fiber sludge and wheat bran media under submerged fermentation. Culture growth conditions were optimized to obtain significant amount of xylanase. Maximum xylanase production was recorded after 72 hours of incubation at 30 °C and 7 pH with 4.0% substrate concentration. In the nutshell, the production of xylanase using inexpensive waste fiber sludge and wheat-bran as an alternative in place of expensive xylan substrate was more cost effective and environment friendly.

Resumo Nos últimos dias, a fonte alternativa de carbono mais barata para fins de fermentação é desejável para minimizar o custo de produção. As xilanases têm se tornado enzimas atraentes como seu potencial no biobranqueamento da indústria de papel e celulose. O objetivo do presente estudo foi identificar a capacidade potencial na produção de xilanase por Bacillus pumilus BS131 isolado localmente usando lodo de fibra residual e farelo de trigo em meio de fermentação submersa. As condições de crescimento da cultura foram otimizadas para obter uma quantidade significativa de xilanase. A produção máxima de xilanase foi registrada após 72 horas de incubação a 30 °C e pH 7 com concentração de substrato de 4,0%. Resumindo, a produção de xilanase usando lodo de fibra residual de baixo custo e farelo de trigo como uma alternativa no lugar do substrato de xilano caro foi mais econômica e ecológica.

Bacillus pumilus proteome changes in response to 2,4,6-trinitrotoluene-induced stress.

2,4,6-Trinitrotoluene (TNT) is the most widely used nitroaromatic compound and is highly resistant to degradation. Most aerobic microorganisms reduce TNT to amino derivatives via formation of nitroso- and hydroxylamine intermediates. Although pathways of TNT degradation are well studied, proteomic analysis of TNT-degrading bacteria was done only for some individual Gram-negative strains. Here, we isolated a Gram-positive strain from TNT-contaminated soil, identified it as Bacillus pumilus using 16S rRNA sequencing, analyzed its growth, the level of TNT transformation, ROS production, and revealed for the first time the bacillary proteome changes at toxic concentration of TNT. The transformation of TNT at all studied concentrations (20-200 mg/L) followed the path of nitro groups reduction with the formation of 4-amino-2,6-dinitrotoluene. Hydrogen peroxide production was detected during TNT transformation. Comparative proteomic analysis of B. pumilus showed that TNT (200 mg/L) inhibited expression of 46 and induced expression of 24 proteins. Among TNT upregulated proteins are those which are responsible for the reductive pathway of xenobiotic transformation, removal of oxidative stress, DNA repair, degradation of RNA and cellular proteins. The production of ribosomal proteins, some important metabolic proteins and proteins involved in cell division are downregulated by this xenobiotic.

An antimicrobial peptide specifically active against Listeria monocytogenes is secreted by Bacillus pumilus SF214.

BACKGROUND: Members of the Bacillus genus produce a large variety of antimicrobial peptides including linear or cyclic lipopeptides and thiopeptides, that often have a broad spectrum of action against Gram-positive and Gram-negative bacteria. We have recently reported that SF214, a marine isolated strain of Bacillus pumilus, produces two different antimicrobials specifically active against either Staphylococcus aureus or Listeria monocytogenes. The anti-Staphylococcus molecule has been previously characterized as a pumilacidin, a nonribosomally synthesized lipopetide composed of a mixture of cyclic heptapeptides linked to fatty acids of variable length. RESULTS: Our analysis on the anti-Listeria molecule of B. pumilus SF214 indicated that it is a peptide slightly smaller than 10 kDa, produced during the exponential phase of growth, stable at a wide range of pH conditions and resistant to various chemical treatments. The peptide showed a lytic activity against growing but not resting cells of Listeria monocytogenes and appeared extremely specific being inactive also against L. innocua, a close relative of L. monocytogenes. CONCLUSIONS: These findings indicate that the B. pumilus peptide is unusual with respect to other antimicrobials both for its time of synthesis and secretion and for its strict specificity against L. monocytogenes. Such specificity, together with its stability, propose this new antimicrobial as a tool for potential biotechnological applications in the fight against the dangerous food-borne pathogen L. monocytogenes.

Design of mutants of GH11 xylanase from Bacillus pumilus for enhanced stability by amino acid substitutions in the N-terminal region: an in silico analysis.

GH11 xylanases are versatile small-molecular-weight single-polypeptide chain monofunctional enzymes. This family of glycoside hydrolases has important applications in food, feed and chemical industries. We designed mutants for improved thermal stability with substitutions in the first six residues of the N-terminal region and evaluated the stability in silico. The first six residues RTITNN of native xylanase have been mutated accordingly to introduce ß structure, increase hydrophobic clusters and enhance conformational rigidity in the molecule. To design stable mutants, the approach consisted of constructing root mean square fluctuation (RMSF) plots of both mesophilic and thermophilic xylanases to check the localized backbone displacement maxima, identify the hydrophobic interaction cluster in and around the peaks of interest, construct mutants by substituting appropriate residues based on beta propensity, hydrophobicity, side chain occupancy and conformational rigidity. This resulted in the decreased number of possible substitutions from 19 to 6 residues. Introduction of conformational rigidity by substitution of asparagine residues at 5th and 6th residue position with proline and valine enhanced the stability. Deletion of N-terminal region increased the stability probably by reducing entropic factors. The structure and stability of GH11 xylanase and resultant mutants were analyzed by root mean square deviation, RMSF, radius of gyration and solvent accessible surface area analysis. The stability of the mutants followed the order N-del > Y1P5 >Y1V5 > ATRLM. The contribution of N-terminal end to overall stability of the molecule is significant because of the proximity of the C-terminal end to the N-terminal end which reinforces long-range interactions. Communicated by Ramaswamy H. Sarma.

Bacillus pumilus induced tolerance of Maize (Zea mays L.) against Cadmium (Cd) stress.

Heavy metals contaminate the soil that alters the properties of soil and negatively affect plants growth. Using microorganism and plant can remove these pollutants from soil. The present investigation was designed to evaluate the induced effect of Bacillus pumilus on maize plant in Cadmium (Cd) contaminated soil. Three different concentrations of Cd (i.e. 0.25, 0.50 and 0.75 mg kg-1) were applied in soil under which maize plants were grown. The germination percentage, shoot length, leaf length, number of leaves, root length, fresh weight and nutrient uptake by maize plant were determined. The experiment was conducted by using complete randomized design (CRD) with three replicates. The result indicated that germination percentage, Shoot length, leaf length, root length, number of leaves, and plant fresh weight were reduced by 37, 39, 39, 32 and 59% respectively at 0.75 mg kg-1 of CdSO4 concentration but when maize seeds inoculated with Bacillus pumilus significantly increased the germination percentage, shoot length, leaf length, number of leaves, plant fresh weight at different concentrations of CdSO4. Moreover, the plant protein were significantly increased by 60% in T6 (0.25 mg kg-1 of CdSO4 + inoculated seed) and Peroxidase dismutase (POD) was also significantly higher by 346% in T6 (0.25 mg kg-1 of CdSO4 + inoculated seed), however, the Superoxide dismutase (SOD) was significantly higher in T5 (0.75 mg kg-1 of CdSO4 + uninoculated seed) and was 769% higher as compared to control. The Cd contents in Bacillus pumilus inoculated maize roots and shoots were decreased. The present investigations indicated that the inoculation of maize plant with Bacillus pumilus can help maize plants to withstand Cd stress but higher concentration of Cd can harm the plant. The Bacillus pumilus has good potential to remediate Cd from soil, and also have potential to reduce the phyto availability and toxicity of Cd.

Modulating the production of xylanase by Bacillus pumilus BS131 through optimization using waste fiber sludge.

In recent days, cheapest alternative carbon source for fermentation purpose is desirable to minimize production cost. Xylanases have become attractive enzymes as their potential in bio-bleaching of pulp and paper industry. The objective of the present study was to identify the potential ability on the xylanase production by locally isolated Bacillus pumilus BS131 by using waste fiber sludge and wheat bran media under submerged fermentation. Culture growth conditions were optimized to obtain significant amount of xylanase. Maximum xylanase production was recorded after 72 hours of incubation at 30 °C and 7 pH with 4.0% substrate concentration. In the nutshell, the production of xylanase using inexpensive waste fiber sludge and wheat-bran as an alternative in place of expensive xylan substrate was more cost effective and environment friendly.

Five Surfactin Isomers Produced during Cheonggukjang Fermentation by Bacillus pumilus HY1 and Their Properties.

The cyclic lipopeptide produced from Bacillus pumilus strain HY1 was isolated from Korean soybean sauce cheonggukjang. The chemical structures of the surfactin isomers were analyzed using matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) and electrospray ionization tandem mass spectrometry (ESI-MS/MS). The five potential surfactin isoforms were detected with protonated masses of m/z 994.7, 1008.7, 1022.7, 1036.7, and 1050.7 and different structures in combination with Na+, K+, and Ca2+ ions. ESI-MS/MS analysis revealed that the isolated surfactin possessed the precise amino acid sequence LLVDLL and hydroxyl fatty acids with 12 to 16 carbons. The surfactin content during cheonggukjang fermentation increased from 0.3 to 51.2 mg/kg over 60 h of fermentation. The mixture of five surfactin isoforms of cheonggukjang inhibited the growth of two cancer cell lines. The growth of both MCF-7 and Caco-2 cells was strongly inhibited with 100 µg/µL of surfactin. This study is the first-time report of five surfactin isomers of Bacillus pumilus strain HY1 during Korean soybean sauce cheonggukjang fermentation, which has cytotoxic properties.

A Novel Microbial Zearalenone Transformation through Phosphorylation.

Zearalenone (ZEA) is a mycotoxin widely occurring in many agricultural commodities. In this study, a purified bacterial isolate, Bacillus sp. S62-W, obtained from one of 104 corn silage samples from various silos located in the United States, exhibited activity to transform the mycotoxin ZEA. A novel microbial transformation product, ZEA-14-phosphate, was detected, purified, and identified by HPLC, LC-MS, and NMR analyses. The isolate has been identified as belonging to the genus Bacillus according to phylogenetic analysis of the 16S rRNA gene and whole genome alignments. The isolate showed high efficacy in transforming ZEA to ZEA-14-phosphate (100% transformation within 24 h) and possessed advantages of acid tolerance (work at pH = 4.0), working under a broad range of temperatures (22-42 °C), and a capability of transforming ZEA at high concentrations (up to 200 µg/mL). In addition, 23 Bacillus strains of various species were tested for their ZEA phosphorylation activity. Thirteen of the Bacillus strains showed phosphorylation functionality at an efficacy of between 20.3% and 99.4% after 24 h incubation, suggesting the metabolism pathway is widely conserved in Bacillus spp. This study established a new transformation system for potential application of controlling ZEA although the metabolism and toxicity of ZEA-14-phosphate requires further investigation.

Biosurfactant from endophytic Bacillus pumilus 2A: physicochemical characterization, production and optimization and potential for plant growth promotion.

BACKGROUND: Microbial surfactants called biosurfactants, thanks to their high biodegradability, low toxicity and stability can be used not only in bioremediation and oil processing, but also in the food and cosmetic industries, and even in medicine. However, the high production costs of microbial surfactants and low efficiency limit their large-scale production. This requires optimization of management conditions, including the possibility of using waste as a carbon source, such as food processing by-products. This papers describes the production and characterization of the biosurfactant obtained from the endophytic bacterial strain Bacillus pumilus 2A grown on various by-products of food processing and its potential applications in supporting plant growth. Four different carbon and nitrogen sources, pH, inoculum concentration and temperature were optimized within Taguchi method. RESULTS: Optimization of bioprocess within Taguchi method and experimental analysis revealed that the optimal conditions for biosurfactant production were brewer's spent grain (5% w/v), ammonium nitrate (1% w/v), pH of 6, 5% of inoculum, and temperature at 30 °C, leading to 6.8 g/L of biosurfactant. Based on gas chromatography-mass spectrometry and Fourier transform infrared spectroscopy analysis produced biosurfactant was determined as glycolipid. Obtained biosurfactant has shown high and long term thermostability, surface tension of 47.7 mN/m, oil displacement of 8 cm and the emulsion index of 69.11%. The examined glycolipid, used in a concentration of 0.2% significantly enhanced growth of Phaseolus vulgaris L. (bean), Raphanus L. (radish), Beta vulgaris L. (beetroot). CONCLUSIONS: The endophytic Bacillus pumilus 2A produce glycolipid biosurfactant with high and long tem thermostability, what makes it useful for many purposes including food processing. The use of brewer's spent grain as the sole carbon source makes the production of biosurfactants profitable, and from an environmental point of view, it is an environmentally friendly way to remove food processing by products. Glycolipid produced by endophytic Bacillus pumilus 2A significantly improve growth of Phaseolus vulgaris L. (bean), Raphanus L. (radish), Beta vulgaris L. (beetroot). Obtained results provide new insight to the possible use of glycolipids as plant growth promoting agents.

Biochemical tests to determine the biodegradability potential of bacterial strains in PAH polluted sites.

Although the use of degrading-bacteria is one of the most efficient methods for the bioremediation of polluted sites, detection, selection and proliferation of the most efficient and competing bacteria is still a challenge. The objective of this multi-stage research was to investigate the effects of the selected bacterial strains on the degradation of anthracene, florentine, naphthalene, and oil, determined by biochemical tests. In the first stage, using the following tests: (a) biosurfactant production (emulsification, oil spreading, number of drops, drop collapse, and surface tension), (b) biofilm production, (c) activity of laccase enzyme, and (d) exopolysaccaride production, the three bacterial strains with the highest degrading potential including Bacillus pumilus, B. aerophilus, and Marinobacter hydrocarbonoclasticus were chosen. In the second stage using the following tests: (a) bacterial growth, (b) laccase enzyme activity, and (c) biosurfactant production (emulsification, oil spreading, and collapse of droplet) the degrading ability of the three selected bacterial strains plus Escherichia coli were compared. Different bacterial strains were able to degrade anthracene, florentine, naphthalene, and oil by the highest rate, three days after inoculation (DAI). However, M. hydrocarbonoclasticus showed the highest rate of florentine degradation. Although with increasing pollutant concentration the degrading potential of the bacterial strains significantly decreased, M. hydrocarbonoclasticus was determined as the most efficient bacterial strain.

Anti-Influenza Activity of the Ribonuclease Binase: Cellular Targets Detected by Quantitative Proteomics.

Unpredictable influenza pandemics, annual epidemics, and sporadic poultry-to-human avian influenza virus infections with high morbidity and mortality rates dictate a need to develop new antiviral approaches. Targeting cellular pathways and processes is a promising antiviral strategy shown to be effective regardless of viral subtypes or viral evolution of drug-resistant variants. Proteomics-based searches provide a tool to reveal the druggable stages of the virus life cycle and to understand the putative antiviral mode of action of the drug(s). Ribonucleases (RNases) of different origins not only demonstrate antiviral effects that are mediated by the direct RNase action on viral and cellular RNAs but can also exert their impact by signal transduction modulation. To our knowledge, studies of the RNase-affected cell proteome have not yet been performed. To reveal cellular targets and explain the mechanisms underlying the antiviral effect employed by the small extra-cellular ribonuclease of Bacillus pumilus (binase) both in vitro and in vivo, qualitative shotgun and quantitative targeted proteomic analyses of the influenza A virus (IAV) H1N1pdm09-infected A549 cells upon binase treatment were performed. We compared proteomes of mock-treated, binase-treated, virus-infected, and virus-infected binase-treated cells to determine the proteins affected by IAV and/or binase. In general, IAV demonstrated a downregulating strategy towards cellular proteins, while binase had an upregulating effect. With the help of bioinformatics approaches, coregulated cellular protein sets were defined and assigned to their biological function; a possible interconnection with the progression of viral infection was conferred. Most of the proteins downregulated by IAV (e.g., AKR1B1, AKR1C1, CCL5, PFN1, RAN, S100A4, etc.) belong to the processes of cellular metabolism, response to stimulus, biological regulation, and cellular localization. Upregulated proteins upon the binase treatment (e.g., AKR1B10, CAP1, HNRNPA2B1, PFN1, PPIA, YWHAB, etc.) are united by the processes of biological regulation, cellular localization, and immune and metabolic processes. The antiviral activity of binase against IAV was expressed by the inversion of virus-induced proteomic changes, resulting in the inhibition of virus-associated processes, including nuclear ribonucleoprotein export (NCL, NPM1, Nup205, and Bax proteins involved) and cytoskeleton remodeling (RDX, PFN1, and TUBB) induced by IAV at the middle stage of single-cycle infection in A549 cells. Modulation of the immune response could be involved as well. Overall, it seems possible that binase exerts its antiviral effects in multiple ways.

Thermostable and alkalistable exopolygalacturonase of Bacillus pumilus dcsr1: Characteristics and applicability.

The bioactive form of thermostable and alkali stable pectinase of Bacillus pumilus dcsr1 is a homodimer of the molecular mass of 60 kDa with a pI of 4.6. The enzyme is optimally active at 50 °C and pH 10.5, and its Michaelis constant (Km), maximum rate of reaction (Vmax), activation energy (Ea), and temperature quotient (Q10) values (for citrus pectin) are 0.29 mg mL-1, 116 µmole mg-1 min-1, 74.73 KJmol-1 and 1.57, respectively. The enzyme has a shelf life of one and a half years at room temperature as well as 4 °C. The activity of the enzyme is stimulated by Mn2+ and Ca2+ and inhibited by Hg+, Cd2+, Co2+, Zn2+, Fe2+, Pb2+, EDTA and urea to a varied extent. The conformational studies of the enzyme revealed a high ß-sheet content in the bioactive dimer, and high α-helix in the inactive monomer. The Circular Dichroism (CD) spectra of the dimer in the presence of inhibitors suggested a marked decrease in ß-sheet, and a significant increase in α-helix, suggesting a key role of ß-sheets in the enzyme catalysis. Based on the end product analysis, the enzyme is an exopolygalacturonase with a unique ability of transglycosylation. When ramie fibers were treated with the enzyme, removal of gummy material (pectin) was visible, confirming its applicability in the degumming process.

Potentiality of living Bacillus pumilus SWU7-1 in biosorption of strontium radionuclide.

Bacillus pumilus SWU7-1 was isolated from strontium ion (Sr(II))-uncontaminated soil, its biosorption potential was evaluated, and the effect of γ-ray radiation treatment on its biosorption was discussed. Domesticated under Sr(II) stress promoted the biosorption ability of B. pumilus to Sr(II), and the biosorption efficiency increased from 46.09% to 94.69%. At a lower initial concentration, the living bacteria had the ability to resist the biosorption of Sr(II). The optimal initial concentration range was 54-130 mg/L. The biosorption profile was better matched by Langmuir than Freundlich model, showing that the biosorption process of Sr(II) by the experimental strain was closer to the surface adsorption. According to Langmuir model, the maximum biosorption capacity of B. pumilus on Sr (II) was 299.4 mg/g. During the bacterial growth in the biosorption process, the changes in biosorption capacity and efficiency can be divided into two phases, and a pseudo-second-order model is followed in each phase. There was no significant difference in the biosorption efficiency of bacteria with different culture time after γ-ray radiation, and all of them were above 90%, which showed that B. pumilus had significant radiation resistance under experimental conditions. This study emphasized the potential application of B. pumilus in the treatment of radioactive Sr(II) pollution by biosorption.

Novel keratinolytic enzymes, discovered from a talented and efficient bacterial keratin degrader.

Huge quantities of keratinaceous waste are a substantial and almost totally unexploited protein resource which could be upgraded for use as high value-added products by efficient keratinolytic enzymes. In this study, we found that Bacillus sp. 8A6 can efficiently degrade chicken feather after 24 h growth. According to phylogenetic analysis, the strain (formerly identified as Bacillus pumilus 8A6) belongs to the B. pumilus species clade but it is more closely related to B. safensis. Hotpep predicted 233 putative proteases from Bacillus sp. 8A6 genome. Proteomic analysis of culture broths from Bacillus sp. 8A6 cultured on chicken feathers or on a mixture of bristles and hooves showed high abundance of proteins with functions related to peptidase activity. Five proteases (one from family M12, one from family S01A, two from family S08A and one from family T3) and four oligopeptide and dipeptide binding proteins were highly expressed when Bacillus sp. 8A6 was grown in keratin media compared to LB medium. This study is the first to report that bacterial proteases in families M12, S01A and T3 are involved in keratin degradation together with proteases from family S08.