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1.
Sheng Wu Gong Cheng Xue Bao ; 37(9): 3242-3252, 2021 Sep 25.
Artigo em Chinês | MEDLINE | ID: mdl-34622632

RESUMO

L-asparaginase hydrolyzes L-asparagine to produce L-aspartic acid and ammonia. It is widely distributed in microorganisms, plants and serum of some rodents, and has important applications in the pharmaceutical and food industries. However, the poor thermal stability, low catalytic efficiency and low yield hampered the further application of L-asparaginase. In this paper, rational design and 5' untranslated region (5'UTR) design strategies were used to increase the specific enzyme activity and protein expression of L-asparaginase derived from Rhizomucor miehei (RmAsnase). The results showed that among the six mutants constructed through homology modeling combined with sequence alignment, the specific enzyme activity of the mutant A344E was 1.5 times higher than the wild type. Subsequently, a food-safe strain Bacillus subtilis 168/pMA5-A344E was constructed, and the UTR strategy was used for the construction of recombinant strain B. subtilis 168/pMA5 UTR-A344E. The enzyme activity of B. subtilis 168/pMA5 UTR-A344E was 7.2 times higher than that of B. subtilis 168/pMA5-A344E. The recombinant strain B. subtilis 168/pMA5 UTR-A344E was scaled up in 5 L fermenter, and the final yield of L-asparaginase was 489.1 U/mL, showing great potential for industrial application.


Assuntos
Asparaginase , Rhizomucor , Asparaginase/biossíntese , Asparaginase/genética , Bacillus subtilis/genética , Microbiologia Industrial , Engenharia de Proteínas , Rhizomucor/enzimologia , Alinhamento de Sequência
2.
Appl Microbiol Biotechnol ; 105(18): 6607-6626, 2021 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-34468804

RESUMO

Bacillus subtilis is a well-characterized Gram-positive bacterium and a valuable host for recombinant protein production because of its efficient secretion ability, high yield, and non-toxicity. Here, we comprehensively review the recent studies on recombinant protein production in B. subtilis to update and supplement other previous reviews. We have focused on several aspects, including optimization of B. subtilis strains, enhancement and regulation of expression, improvement of secretion level, surface display of proteins, and fermentation optimization. Among them, optimization of B. subtilis strains mainly involves undirected chemical/physical mutagenesis and selection and genetic manipulation; enhancement and regulation of expression comprises autonomous plasmid and integrated expression, promoter regulation and engineering, and fine-tuning gene expression based on proteases and molecular chaperones; improvement of secretion level predominantly involves secretion pathway and signal peptide screening and optimization; surface display of proteins includes surface display of proteins on spores or vegetative cells; and fermentation optimization incorporates medium optimization, process condition optimization, and feeding strategy optimization. Furthermore, we propose some novel methods and future challenges for recombinant protein production in B. subtilis.Key points• A comprehensive review on recombinant protein production in Bacillus subtilis.• Novel techniques facilitate recombinant protein expression and secretion.• Surface display of proteins has significant potential for different applications.


Assuntos
Bacillus subtilis , Proteínas de Bactérias , Bacillus subtilis/genética , Proteínas de Bactérias/genética , Chaperonas Moleculares , Sinais Direcionadores de Proteínas , Proteínas Recombinantes/genética
3.
Appl Microbiol Biotechnol ; 105(18): 6853-6870, 2021 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-34477941

RESUMO

In wild strains of Bacillus, a handful of extracellular natural products act as signals that can regulate multicellular behavior, but relatively little is known about molecular mechanisms' detail. We proposed a previously unreported molecular mechanism for triggering multicellularity in B. velezensis Bs916 by an endogenous cyclic lipopeptide, bacillomycin L. The genome-wide effect on gene expression was caused by the disruption of bacillomycin L gene cluster, and 100 µg/mL bacillomycin L was revealed by quantitative transcriptomics. A total of 878 differentially expressed genes among Bs916, Δbl, and Δbl + 100BL were identified and grouped into 9 functional categories. The transcription levels of 40 candidate genes were further evaluated by RT-qPCR analysis. The expression of eight candidate genes regulated by bacillomycin L in a dose-dependent manner was revealed by LacZ fusion experiment. Although the addition of bacillomycin L could not completely restore the expression levels of the differentially regulated genes in △bl, our results strongly suggest that bacillomycin L acts as a tuning signal of swarming motility and complex biofilm formation by indirectly regulating the expression levels of some two-component systems (TCSs) connector genes, particularly including several Raps that potentially regulate the phosphorylation levels of three major regulators ComA, DegU, and Spo0A.Key points• Proposed model for bacillomycin L regulation in B. velezensis Bs916.• Bacillomycin L can act as an extracellular signal to regulate the phosphorylation levels of three major regulators, ComA, DegU, and Spo0A and control the multicellular processes of vegetative growth, competent, motility, matrix production, sporulation, and autolysis.


Assuntos
Bacillus , Lipopeptídeos , Peptídeos Catiônicos Antimicrobianos , Bacillus/genética , Bacillus subtilis , Peptídeos Cíclicos
4.
Enzyme Microb Technol ; 150: 109863, 2021 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-34489022

RESUMO

Protein-glutaminase (EC 3.5.1.44, PG) converts protein glutamine residues in proteins and peptides into glutamic acid residue, and markedly improves the solubility, emulsification, and foaming properties of food proteins. However, the source bacteria, Chryseobacterium proteolyticum, have low enzyme production ability, inefficient genetic operation, and high production cost. Therefore, it is critical to establish an efficient expression system for active PG. Here, combinatorial engineering was developed for high-yield production of PG in Bacillus subtilis. First, we evaluated different B. subtilis strains for PG self-activation. Then, combinatorial optimization involving promoters, signal peptides, and culture medium was applied to produce active recombinant PG in a B. subtilis expression system. Through combinatorial engineering, PG enzyme activity reached 3.23 U/mL in shaken-flask cultures. Active PG with the yield of 7.07 U/mL was obtained at 40 h by the PSecA-YdeJ combination in fed-batch fermentation, which is the highest yield of PG in existing reports.


Assuntos
Bacillus subtilis , Proteínas de Bactérias/biossíntese , Chryseobacterium , Glutaminase/biossíntese , Bacillus subtilis/metabolismo , Chryseobacterium/enzimologia , Fermentação , Engenharia de Proteínas , Sinais Direcionadores de Proteínas
5.
Sheng Wu Gong Cheng Xue Bao ; 37(8): 2825-2835, 2021 Aug 25.
Artigo em Chinês | MEDLINE | ID: mdl-34472300

RESUMO

Biosynthesis of nanomaterials has attracted much attention for its excellent characteristics such as low energy consumption, high safety, and environmental friendliness. As we all know, the toxic selenite can be transformed into higher-value nanomaterials by using bacteria. In this study, nano-selenium was synthesized by halophilic Bacillus subtilis subspecies stercoris strain XP in LB medium supplemented with selenite (electron acceptor). The physicochemical characteristics of nano-selenium were analyzed by scanning electron microscope (SEM), X-ray energy dispersive spectral analysis (EDAX), X-ray diffraction (XRD), and fourier transform infrared spectroscopy (FTIR). Meanwhile, the antifungal activity of nano-selenium to strawberry pathogens (fusarium wilt, erythema, and purple spot fungi) was determined. The products from reduction of selenite by strain XP was amorphous spherical selenium nanoparticles (SeNPs) with a diameter range of 135-165 nm. The production of SeNPs was positively correlated with time (0-48 h) and no changes were observed on cell morphology. Selenium was dominant in the surface of SeNPs where the organic elements (C, O, N, and S) existed at the same time. SeNPs were coated with biomolecules containing functional groups (such as -OH, C=O, N-H, and C-H) which were associated with the stability and bioactivity of particles. Although the highest concentration of SeNPs had significant (P<0.05) inhibitory effects on three strains of strawberry pathogens, antifungal activity to erythema and fusarium wilt pathogenic fungi was higher than that to purple spot pathogenic fungi from strawberry. In conclusion, strain XP not only has strong tolerance to high salt stress, but can be also used to synthesize biological SeNPs with good stability and biological activity. Thus, the strain XP has bright perspectives and great potential advantage in pathogens control and green selenium-rich strawberry planting as well as other fields.


Assuntos
Fragaria , Nanopartículas , Selênio , Bacillus subtilis , Ácido Selenioso
6.
Nat Commun ; 12(1): 5429, 2021 09 14.
Artigo em Inglês | MEDLINE | ID: mdl-34521822

RESUMO

Bacillus subtilis is a model gram-positive bacterium, commonly used to explore questions across bacterial cell biology and for industrial uses. To enable greater understanding and control of proteins in B. subtilis, here we report broad and efficient genetic code expansion in B. subtilis by incorporating 20 distinct non-standard amino acids within proteins using 3 different families of genetic code expansion systems and two choices of codons. We use these systems to achieve click-labelling, photo-crosslinking, and translational titration. These tools allow us to demonstrate differences between E. coli and B. subtilis stop codon suppression, validate a predicted protein-protein binding interface, and begin to interrogate properties underlying bacterial cytokinesis by precisely modulating cell division dynamics in vivo. We expect that the establishment of this simple and easily accessible chemical biology system in B. subtilis will help uncover an abundance of biological insights and aid genetic code expansion in other organisms.


Assuntos
Aminoácidos/genética , Aminoacil-tRNA Sintetases/genética , Bacillus subtilis/genética , Proteínas de Bactérias/genética , Regulação Bacteriana da Expressão Gênica , Código Genético , Aminoácidos/química , Aminoácidos/metabolismo , Aminoacil-tRNA Sintetases/classificação , Aminoacil-tRNA Sintetases/metabolismo , Bacillus subtilis/metabolismo , Proteínas de Bactérias/metabolismo , Códon , Citocinese/genética , Escherichia coli/genética , Escherichia coli/metabolismo , Genoma Bacteriano , Ligação Proteica , Biossíntese de Proteínas , Mapeamento de Interação de Proteínas , RNA de Transferência/genética , RNA de Transferência/metabolismo
7.
J Agric Food Chem ; 69(38): 11272-11281, 2021 Sep 29.
Artigo em Inglês | MEDLINE | ID: mdl-34546731

RESUMO

The European Union banned the usage of antibiotic growth promoters in animal production. The probiotic microorganism of the genus Bacillus appeared to be an attractive candidate to replace antibiotics. The Bacillus subtilis DSM 29784 is one of these strains. To date, the probiotic effect has not been completely understood, but it is supposed that the effect depends on metabolites of the microorganism. Imaging high-performance thin-layer chromatography (HPTLC) is a powerful tool to visualize differences in the metabolite profile of bacteria with high genetic similarity to allow a better understanding of the probiotic effect. In comparison to other bacteria, especially these bacterial cells were more robust to harsh cultivation conditions and produced a higher level of antioxidants or bioactive substances such as surfactin. HPTLC enabled the comparison of pure cell cultures to the spore cultivation in the feed, and the results explain and support the probiotic effect.


Assuntos
Bacillus , Probióticos , Ração Animal , Animais , Antibacterianos , Bacillus subtilis , Cromatografia em Camada Delgada
8.
Curr Microbiol ; 78(11): 3968-3979, 2021 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-34550433

RESUMO

In this study, a total 30 rhizobacterial isolates were screened out based on resistance against different concentrations of mercuric chloride (HgCl2), growth on nitrogen-free mannitol (NFM) and production of indole-3-acetic acid (IAA). The biochemical and plant growth promoting characterization of selected isolates was performed by different biochemical tests. Out of 30, six isolates, UM-3, AZ-5, UM-7, UM-11, UM-26, and UM-28 showed resistance at 30 µg/ml HgCl2, pronounced growth on NFM and high production of IAA as 18.6, 16.7, 16, 18.7, 14, and 16 µg/ml, respectively (P < 0.05). The 16S rDNA ribotyping and phylogenetic analysis of selected bacterial isolates were performed and characterized as Exiguobacterium sp. UM-3 (KJ736011), Bacillus thuringiensis AZ-5 (KJ675627), Bacillus subtilis UM-7 (KJ736013), Enterobacter cloacae UM-11 (KJ736014), Pseudomonas aeruginosa UM-26 (KJ736016), P. aeruginosa UM-28 (KJ736017) and Bacillus pumilus UM-16 (KJ736015) used as negative control. B. thuringiensis AZ-5 showed high resistance against 30 µg/ml of HgCl2 due to the presence of merB gene. The structural determination of MerB protein was carried out using bioinformatics tools, i.e., Protparam, Pfam, InterProScan, STRING, Jpred4, PSIPRED, I-TASSER, COACH server and ERRAT. These tools predicted the structural based functional homology of MerB protein (organomercuric lyase) in association with MerA (mercuric reductase) in bacterial Hg-detoxification system.


Assuntos
Mercúrio , Bacillus subtilis , Simulação por Computador , Filogenia , Desenvolvimento Vegetal
9.
J Anim Sci ; 99(10)2021 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-34473279

RESUMO

The present study used intrauterine growth restriction (IUGR) piglets as an animal model to determine the effect of Bacillus subtilis on intestinal integrity, antioxidant capacity, and microbiota in the jejunum of suckling piglets. In total, 8 normal birth weight (NBW) newborn piglets (1.62 ± 0.10 kg) and 16 newborn IUGR piglets (0.90 ± 0.08 kg) were selected and assigned to three groups. Piglets were orally gavaged with 10-mL sterile saline (NBW and IUGR groups), and IUGR piglets were orally gavaged with 10-mL/d bacterial fluid (B. subtilis diluted in sterile saline, gavage in the dose of 2 × 109 colony-forming units per kg of body weight; IBS group; n = 8). IUGR induced jejunal barrier dysfunction and redox status imbalance of piglets, and changed the abundances of bacteria in the jejunum. Treatment with B. subtilis increased (P < 0.05) the ratio of villus height to crypt depth (VH/CD) in the jejunum, decreased (P < 0.05) the plasma diamine oxidase (DAO) activity, and enhanced (P < 0.05) the gene expressions of zonula occludens-1 (ZO-1), occludin, and claudin-1 in the jejunum of IUGR piglets. Treatment with B. subtilis decreased (P < 0.05) the concentration of protein carbonyl (PC) and increased (P < 0.05) the activities of catalase (CAT) and total superoxide dismutase (T-SOD) in the jejunum of IUGR piglets. Treatment with B. subtilis also increased (P < 0.05) gene expressions of superoxide dismutase 1 (SOD1), CAT, and nuclear factor erythroid 2-related factor (Nrf2), as well as the protein expressions of heme oxygenase-1 (HO-1), SOD1, and Nrf2 in the jejunum of IUGR piglets. Treatment with B. subtilis also improved the abundances and the community structure of bacteria in the jejunum of IUGR piglets. These results suggested that IUGR damaged the jejunal barrier function and antioxidant capacity of suckling piglets, and altered the abundances of bacteria in the jejunum. Treatment with B. subtilis improved the intestinal integrity and antioxidant capacity while also improved the abundances and structure of bacteria in the jejunum of suckling piglets.


Assuntos
Retardo do Crescimento Fetal , Doenças dos Suínos , Animais , Bacillus subtilis , Retardo do Crescimento Fetal/veterinária , Jejuno , Oxirredução , Suínos
10.
Bioresour Technol ; 341: 125836, 2021 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-34469820

RESUMO

Diacetylchitobiose deacetylase (Dac) from Pyrococcus horikoshii can realize the one-step production of glucosamine (GlcN). The efficient expression and secretion of Dac play a central role in the green production of GlcN. In this study, Bacillus subtilis WB600 was used as the expression host. Firstly, we screened 12 signal peptides, among which signal peptide NprB had the strongest ability of guiding Dac secretion. Further optimization of the functional region showed that the extracellular Dac activity of NprB mutant was increased to 3682.2 U/mL. Next, the extracellular Dac activity was increased to 4807.6 U/mL by RBS sequence optimization. Then we got a new recombinant B. subtilis C6 for plasmid-free expression of Dac by integrating comK gene and silencing bpr, nprB, aprE, mpr and nprE genes. Finally, the extracellular Dac activity of genome-integrating strain reached 6357.38 U/mL, which was the highest level reported so far.


Assuntos
Bacillus subtilis , Glucosamina , Bacillus subtilis/genética , Proteínas de Bactérias/genética , Plasmídeos/genética
11.
Biochemistry (Mosc) ; 86(8): 942-951, 2021 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-34488571

RESUMO

Translation of the genetic information into proteins, performed by the ribosome, is a key cellular process in all organisms. Translation usually proceeds smoothly, but, unfortunately, undesirable events can lead to stalling of translating ribosomes. To rescue these faulty arrested ribosomes, bacterial cells possess three well-characterized quality control systems, tmRNA, ArfA, and ArfB. Recently, an additional ribosome rescue mechanism has been discovered in Bacillus subtilis. In contrast to the "canonical" systems targeting the 70S bacterial ribosome, this latter mechanism operates by first splitting the ribosome into the small (30S) and large (50S) subunits to then clearing the resultant jammed large subunit from the incomplete nascent polypeptide. Here, I will discuss the recent microbiological, biochemical, and structural data regarding functioning of this novel rescue system.


Assuntos
Biossíntese de Proteínas , RNA Bacteriano/química , Ribossomos/química , Bacillus subtilis/metabolismo , Proteínas de Bactérias/genética , Bioquímica , Escherichia coli/metabolismo , Proteínas de Escherichia coli/genética , Conformação de Ácido Nucleico , Peptídeos/química , RNA Mensageiro/metabolismo , Proteínas de Ligação a RNA/metabolismo
12.
Biochemistry (Mosc) ; 86(8): 962-975, 2021 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-34488573

RESUMO

Discovered almost twenty years ago, riboswitches turned out to be one of the most common regulatory systems in bacteria, with representatives found in eukaryotes and archaea. Unlike many other regulatory elements, riboswitches are entirely composed of RNA and capable of modulating expression of genes by direct binding of small cellular molecules. While bacterial riboswitches had been initially thought to control production of enzymes and transporters associated with small organic molecules via feedback regulatory circuits, later findings identified riboswitches directing expression of a wide range of genes and responding to various classes of molecules, including ions, signaling molecules, and others. The 5'-untranslated mRNA regions host a vast majority of riboswitches, which modulate transcription or translation of downstream genes through conformational rearrangements in the ligand-sensing domains and adjacent expression-controlling platforms. Over years, the repertoire of regulatory mechanisms employed by riboswitches has greatly expanded; most recent studies have highlighted the importance of alternative mechanisms, such as RNA degradation, for the riboswitch-mediated genetic circuits. This review discusses the plethora of bacterial riboswitch mechanisms and illustrates how riboswitches utilize different features and approaches to elicit various regulatory responses.


Assuntos
Estabilidade de RNA , Riboswitch/fisiologia , Regiões 5' não Traduzidas , Bacillus subtilis , Bactérias/metabolismo , Escherichia coli , Regulação Bacteriana da Expressão Gênica , Ligantes , Fases de Leitura Aberta , RNA/metabolismo , RNA Mensageiro/metabolismo , Transdução de Sinais
13.
Front Cell Infect Microbiol ; 11: 722778, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34557426

RESUMO

Beneficial and probiotic bacteria play an important role in conferring immunity of their hosts to a wide range of bacterial, viral, and fungal diseases. Bacillus subtilis is a Gram-positive bacterium that protects the plant from various pathogens due to its capacity to produce an extensive repertoire of antibiotics. At the same time, the plant microbiome is a highly competitive niche, with multiple microbial species competing for space and resources, a competition that can be determined by the antagonistic potential of each microbiome member. Therefore, regulating antibiotic production in the rhizosphere is of great importance for the elimination of pathogens and establishing beneficial host-associated communities. In this work, we used B. subtilis as a model to investigate the role of plant colonization in antibiotic production. Flow cytometry and imaging flow cytometry (IFC) analysis supported the notion that Arabidopsis thaliana specifically induced the transcription of the biosynthetic clusters for the non-ribosomal peptides surfactin, bacilysin, plipastatin, and the polyketide bacillaene. IFC was more robust in quantifying the inducing effects of A. thaliana, considering the overall heterogeneity of the population. Our results highlight IFC as a useful tool to study the effect of association with a plant host on bacterial gene expression. Furthermore, the common regulation of multiple biosynthetic clusters for antibiotic production by the plant can be translated to improve the performance and competitiveness of beneficial members of the plant microbiome.


Assuntos
Arabidopsis , Bacillus subtilis , Antibacterianos , Bacillus subtilis/genética , Genes Bacterianos , Raízes de Plantas
14.
J Agric Food Chem ; 69(40): 11835-11846, 2021 Oct 13.
Artigo em Inglês | MEDLINE | ID: mdl-34590486

RESUMO

BsCsn46A, a GH46 family chitosanase from Bacillus subtilis, has great potential for industrial chitooligosaccharide production due to its high activity and stability. In this study, a special amino acid Pro121 was identified not fit in the helix structure, which was located in the opposite side of the active center in BsCsn46A, by the PoPMuSiC algorithm. Then, saturation mutagenesis was performed to explore the role of the site amino acid 121. Compared with the wild type, the specific activity of P121N, P121C, and P121V was increased by 1.69-, 1.97-, and 2.15-fold, respectively. In particular, the specific activity of P121N was increased without loss of thermostability, indicating that replacing the structural stiffness of proline in the helical structure could significantly improve the chitosanase activity. The Km values of P121N, P121C, and P121V decreased significantly, indicating that the affinity between the enzyme-substrate complex was enhanced. Through molecular docking, it was found that the increase of hydrogen bonds and van der Waals force between the enzyme-substrate complex and the removal of unfavorable bonds might be the main reason for the change of enzyme properties. In addition, the optimal temperature of the three mutants changed from 60 to 55 °C. These results indicate that the site 121 plays a critical role in the catalytic activity and enzymatic properties of chitosanase. To our knowledge, the results provide novel data on chitosanase activity and identify an excellent candidate of industrial chitosanase.


Assuntos
Bacillus subtilis , Glicosídeo Hidrolases , Bacillus subtilis/genética , Bacillus subtilis/metabolismo , Estabilidade Enzimática , Glicosídeo Hidrolases/genética , Glicosídeo Hidrolases/metabolismo , Simulação de Acoplamento Molecular , Mutagênese , Temperatura
15.
J Agric Food Chem ; 69(35): 10184-10193, 2021 Sep 08.
Artigo em Inglês | MEDLINE | ID: mdl-34449206

RESUMO

The flavanoid hesperidin (Hsd) is one of the major polyphenols in citrus fruits. Hsd and its aglycone hesperetin (Hst) have a broad array of bioactivities; however, their low aqueous solubility and low intestinal permeability lead to their limited oral bioavailability. In the present study, we generated two water-soluble derivatives of Hst, namely, Hst 7-O-phosphate and Hst3'-O-phosphate, by a unique bioconversion process of Bacillus subtilis var. natto BCRC80517. The phosphorylated products showed superior aqueous solubility and distinct physicochemical properties compared with the original Hst. The Hst phosphate derivatives (HstPs) remained stable in simulated gastric and intestinal fluids for 240 min and could revert to the original Hst form by alkaline phosphatase treatment in Caco-2 cells, showing enhanced intestinal permeability in vitro. After oral administration in rats, HstPs greatly elevated plasma exposure to Hst and showed better bioavailability than did Hsd. HstPs may be a potential and efficient alternative to Hst.


Assuntos
Hesperidina , Animais , Bacillus subtilis , Disponibilidade Biológica , Células CACO-2 , Humanos , Fosforilação , Ratos
16.
J Med Food ; 24(8): 860-865, 2021 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-34406879

RESUMO

Bacillus subtilis fermented soybeans (natto) contain high vitamin K2 levels, mostly as menaquinone-7 (MK-7), and must be avoided by warfarin-dependent patients. This is the first report which demonstrates the characteristics and clinical relevance of a low MK-7 natto for such patients. We generated a novel, mutant B. subtilis strain TTCC2051 with short-term fermentation and reduced MK-7 production, yielding 19-24% of the normal MK-7 content. After functional assessments and a preclinical trial, 10 warfarin-dependent patients underwent a clinical trial with a 7-day ingestion test of the low MK-7 natto. Functional assessments were satisfactory, and the preclinical trial showed no increases in plasma MK-7 levels after 7 days of ingestion. In the clinical trial, 20 g/day of the low MK-7 natto significantly increased plasma MK-7 levels while 10 g/day did not. However, neither dose of low MK-7 natto changed international normalized ratio of prothrombin time (PT-INR) values in either group. The low MK-7 natto neither changed PT-INR values nor precipitated adverse events if ingested with a once-daily maximum of 20 g (46 µg of MK-7). Thus, this novel food product has potential for consumption by warfarin-dependent patients.


Assuntos
Alimentos de Soja , Varfarina , Bacillus subtilis , Fermentação , Humanos , Soja , Vitamina K 2/análogos & derivados
17.
Poult Sci ; 100(9): 101372, 2021 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-34364120

RESUMO

The global poultry trend toward the more responsible use of antibiotics is becoming recurrent and has demanded the need to generate new natural alternatives. Probiotics have gained importance as an option to use as growth promoters. This study aimed to evaluate Bacillus subtillis QST713 as a substitute for an antibiotic growth promoter (BMD). A total of 150 male broilers were assigned to three dietary treatments: 1) control diet (CO), 2) control diet + 500 g/t of BMD (AGP), and 3) control diet + 100 g/t of B. subtilis QST713 (PB), respectively. Each treatment was monitored for 5 wk for the productive variables: body weight, accumulated feed consumption, food conversion, and European efficiency factor. At the end of each week, fresh fecal samples were cultured and quantified for E. coli, Enterococcus spp., and Lactobacillus spp. At the end of the trial, blood samples were analyzed for hemogram and intestinal samples (anterior portion) for histomorphometry. The data were statistically analyzed with an analysis of variance and subjected to a least significant difference test (Tukey). The zootechnical yields were similar in the AGP and PB groups (P ˃ 0.05); both superior to the control group. In the hematological profiles, no difference was observed between the experimental groups. E. coli and Enterococcus counts were significantly lower (P ˂ 0.05), and Lactobacillus counts were significantly (P ˂ 0.05) higher in the PB group, relative to CO and AGP groups. No differences (P ˃ 0.05) were found in bacterial counts between the CO and AGP groups. The intestinal mucosa and villi in the PB group were significantly (P ˂ 0.05) longer and with less deeper crypts than CO and AGP groups. We conclude that B. subtillis QST713, used at the suggested commercial dose (100 g/ton), is an effective growth-promoting alternative to BMD that modulates the microbiota and intestinal architecture, thus producing zootechnical yields consistent with BMD.


Assuntos
Bacitracina , Probióticos , Ração Animal/análise , Animais , Bacillus subtilis , Bacitracina/farmacologia , Galinhas , Dieta/veterinária , Escherichia coli , Masculino
18.
Pestic Biochem Physiol ; 178: 104916, 2021 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-34446192

RESUMO

Soybean root rot occurs globally and seriously affects soybean production. To avoid the many disadvantages of chemical fungicides, the addition of Bacillus is gradually becoming an alternative strategy to tackle soybean root rot. However, the molecular mechanism of phytopathogenic fungi in this process by Bacillus inhibition is rarely reported. In this study, we isolated a strain of B. subtilis HSY21 from soybean rhizosphere soil, which had an inhibition rate of 81.30 ± 0.15% (P < 0.05) against Fusarium oxysporum. The control effects of this strain against soybean root rot under greenhouse and field conditions were 63.83% and 57.07% (P < 0.05), respectively. RNA-seq analysis of F. oxysporum after treatment with strain HSY21 revealed 1445 downregulated genes and 1561 upregulated genes. Among them, genes involved in mycelial growth, metabolism regulation, and disease-related enzymes were mostly downregulated. The activities of cellulase, ß-glucosidase, α-amylase, and pectin-methyl- galacturonase as well as levels of oxalic acid and ergosterol in F. oxysporum were significantly decreased after HSY21 treatment. These results demonstrated that B. subtilis HSY21 could effectively control F. oxysporum by inhibiting its growth and the expression of pathogenic genes, thus indicating that this strain may be an ideal candidate for the prevention and control of soybean root rot.


Assuntos
Fusarium , Bacillus subtilis/genética , Fusarium/genética , Doenças das Plantas/genética , Soja/genética , Virulência
19.
Mol Cell ; 81(16): 3310-3322.e6, 2021 08 19.
Artigo em Inglês | MEDLINE | ID: mdl-34416138

RESUMO

Amino acid starvation is sensed by Escherichia coli RelA and Bacillus subtilis Rel through monitoring the aminoacylation status of ribosomal A-site tRNA. These enzymes are positively regulated by their product-the alarmone nucleotide (p)ppGpp-through an unknown mechanism. The (p)ppGpp-synthetic activity of Rel/RelA is controlled via auto-inhibition by the hydrolase/pseudo-hydrolase (HD/pseudo-HD) domain within the enzymatic N-terminal domain region (NTD). We localize the allosteric pppGpp site to the interface between the SYNTH and pseudo-HD/HD domains, with the alarmone stimulating Rel/RelA by exploiting intra-NTD autoinhibition dynamics. We show that without stimulation by pppGpp, starved ribosomes cannot efficiently activate Rel/RelA. Compromised activation by pppGpp ablates Rel/RelA function in vivo, suggesting that regulation by the second messenger (p)ppGpp is necessary for mounting an acute starvation response via coordinated enzymatic activity of individual Rel/RelA molecules. Control by (p)ppGpp is lacking in the E. coli (p)ppGpp synthetase SpoT, thus explaining its weak synthetase activity.


Assuntos
Regulação Alostérica/genética , Proteínas de Escherichia coli/genética , GTP Pirofosfoquinase/genética , Guanosina Pentafosfato/genética , Pirofosfatases/genética , Aminoácidos/metabolismo , Bacillus subtilis/genética , Bacillus subtilis/metabolismo , Domínio Catalítico/genética , Escherichia coli/genética , Escherichia coli/metabolismo , Hidrolases/genética , Ribossomos/genética , Ribossomos/metabolismo , Inanição/genética , Inanição/metabolismo
20.
Nat Microbiol ; 6(9): 1175-1187, 2021 09.
Artigo em Inglês | MEDLINE | ID: mdl-34373624

RESUMO

Most bacteria replicate and segregate their DNA concomitantly while growing, before cell division takes place. How bacteria synchronize these different cell cycle events to ensure faithful chromosome inheritance by daughter cells is poorly understood. Here, we identify Cell Cycle Regulator protein interacting with FtsZ (CcrZ) as a conserved and essential protein in pneumococci and related Firmicutes such as Bacillus subtilis and Staphylococcus aureus. CcrZ couples cell division with DNA replication by controlling the activity of the master initiator of DNA replication, DnaA. The absence of CcrZ causes mis-timed and reduced initiation of DNA replication, which subsequently results in aberrant cell division. We show that CcrZ from Streptococcus pneumoniae interacts directly with the cytoskeleton protein FtsZ, which places CcrZ in the middle of the newborn cell where the DnaA-bound origin is positioned. This work uncovers a mechanism for control of the bacterial cell cycle in which CcrZ controls DnaA activity to ensure that the chromosome is replicated at the right time during the cell cycle.


Assuntos
Proteínas de Bactérias/metabolismo , Ciclo Celular , Proteínas do Citoesqueleto/metabolismo , Replicação do DNA , Streptococcus pneumoniae/citologia , Streptococcus pneumoniae/metabolismo , Bacillus subtilis/citologia , Bacillus subtilis/genética , Bacillus subtilis/metabolismo , Proteínas de Bactérias/genética , Proteínas do Citoesqueleto/genética , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Ligação Proteica , Streptococcus pneumoniae/genética
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