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1.
Molecules ; 27(22)2022 Nov 17.
Artigo em Inglês | MEDLINE | ID: mdl-36432083

RESUMO

In an effort to explore bioactive anti-inflammatory compounds from natural Actinobacteria resources from all over Taiwan and various ecological environments, an active strain of Acrocarpospora punica was collected at Taitung County in Taiwan, prepared from soil origin. A bioassay-guided fractionation of the BuOH extract of a culture broth of a new strain of the actinomycete Acrocarpospora punica led to the isolation of five previously undescribed compounds: acrocarpunicains A-F (1-6). The structures were elucidated by 1D and 2D Nuclear Magnetic Resonance (NMR) spectroscopy and mass spectrometry. Furthermore, the isolated compounds were subjected to in vitro testing to evaluate their anti-inflammatory activity. Of these isolates, acrocarpunicains A (1), B (2), C (3) and F (6) showed NO inhibitory activity with IC50 values of 9.36 ± 0.25, 10.11 ± 0.47, 5.15 ± 0.18, and 27.17 ± 1.87 µM, stronger than the positive control, quercetin (IC50 = 35.95 ± 2.34 µM). To the best of our knowledge, this is the first report on azaphilone and phenanthrene-type metabolites from the genus Acrocarpospora.


Assuntos
Actinobacteria , Actinomycetales , Actinobacteria/metabolismo , Actinomycetales/química , Anti-Inflamatórios , Espectroscopia de Ressonância Magnética , Taiwan
2.
Molecules ; 27(21)2022 Oct 27.
Artigo em Inglês | MEDLINE | ID: mdl-36364123

RESUMO

Periodontitis, as one of the most common diseases on a global scale, is a public health concern. Microbial resistance to currently available antimicrobial agents is becoming a growing issue in periodontal treatment. As a result, it is critical to develop effective and environmentally friendly biomedical approaches to overcome such challenges. The investigation of Streptomyces rochei MS-37's performance may be the first of its kind as a novel marine actinobacterium for the green biosynthesis of silver nanoparticles (SNPs) and potentials as antibacterial, anti-inflammatory, antibiofilm, and antioxidant candidates suppressing membrane-associated dental infections. Streptomyces rochei MS-37, a new marine actinobacterial strain, was used in this study for the biosynthesis of silver nanoparticles for various biomedical applications. Surface plasmon resonance spectroscopy showed a peak at 429 nm for the SNPs. The SNPs were spherical, tiny (average 23.2 nm by TEM, 59.4 nm by DLS), very stable (-26 mV), and contained capping agents. The minimum inhibitory concentrations of the SNPs that showed potential antibacterial action ranged from 8 to 128 µg/mL. Periodontal pathogens were used to perform qualitative evaluations of microbial adhesion and bacterial penetration through guided tissue regeneration membranes. The findings suggested that the presence of the SNPs could aid in the suppression of membrane-associated infection. Furthermore, when the anti-inflammatory action of the SNPs was tested using nitric oxide radical scavenging capacity and protein denaturation inhibition, it was discovered that the SNPs were extremely efficient at scavenging nitric oxide free radicals and had a strong anti-denaturation impact. The SNPs were found to be more cytotoxic to CAL27 than to human peripheral blood mononuclear cells (PBMCs), with IC50 values of 81.16 µg/mL in PBMCs and 34.03 µg/mL in CAL27. This study's findings open a new avenue for using marine actinobacteria for silver nanoparticle biosynthesis, which holds great promise for a variety of biomedical applications, in particular periodontal treatment.


Assuntos
Actinobacteria , Nanopartículas Metálicas , Streptomyces , Humanos , Prata/química , Nanopartículas Metálicas/química , Leucócitos Mononucleares/metabolismo , Streptomyces/metabolismo , Testes de Sensibilidade Microbiana , Antibacterianos/química , Actinobacteria/metabolismo , Extratos Vegetais/química , Espectroscopia de Infravermelho com Transformada de Fourier
3.
Appl Environ Microbiol ; 88(20): e0133722, 2022 Oct 26.
Artigo em Inglês | MEDLINE | ID: mdl-36190258

RESUMO

Two-component systems (TCSs) act as common regulatory systems allowing bacteria to detect and respond to multiple environmental stimuli, including cell envelope stress. The MtrAB TCS of Actinobacteria is critical for cell wall homeostasis, cell proliferation, osmoprotection, and antibiotic resistance, and thus is found to be highly conserved across this phylum. However, how precisely the MtrAB TCS regulates cellular homeostasis in response to environmental stress remains unclear. Here, we show that the MtrAB TCS plays an important role in the tolerance to different types of cell envelope stresses, including environmental stresses (i.e., oxidative stress, lysozyme, SDS, osmotic pressure, and alkaline pH stresses) and envelope-targeting antibiotics (i.e., isoniazid, ethambutol, glycopeptide, and ß-lactam antibiotics) in Dietzia sp. DQ12-45-1b. An mtrAB mutant strain exhibited slower growth compared to the wild-type strain and was characterized by abnormal cell shapes when exposed to various environmental stresses. Moreover, deletion of mtrAB resulted in decreased resistance to isoniazid, ethambutol, and ß-lactam antibiotics. Further, Cleavage under targets and tagmentation sequencing (CUT&Tag-seq) and electrophoretic mobility shift assays (EMSAs) revealed that MtrA binds the promoters of genes involved in peptidoglycan biosynthesis (ldtB, ldtA, murJ), hydrolysis (GJR88_03483, GJR88_4713), and cell division (ftsE). Together, our findings demonstrated that the MtrAB TCS is essential for the survival of Dietzia sp. DQ12-45-1b under various cell envelope stresses, primarily by controlling multiple downstream cellular pathways. Our work suggests that TCSs act as global sensors and regulators in maintaining cellular homeostasis, such as during episodes of various environmental stresses. The present study should shed light on the understanding of mechanisms for bacterial adaptivity to extreme environments. IMPORTANCE The multilayered cell envelope is the first line of bacterial defense against various extreme environments. Bacteria utilize a large number of sensing and regulatory systems to maintain cell envelope homeostasis under multiple stress conditions. The two-component system (TCS) is the main sensing and responding apparatus for environmental adaptation. The MtrAB TCS highly conserved in Actinobacteria is critical for cell wall homeostasis, cell proliferation, osmoprotection, and antibiotic resistance. However, how MtrAB works with regard to signals impacting changes to the cell envelope is not fully understood. Here, we found that in the Actinobacterium Dietzia sp. DQ12-45-1b, a TCS named MtrAB is pivotal for ensuring normal cell growth as well as maintaining proper cell morphology in response to various cell envelope stresses, namely, by regulating the expression of cell envelope-related genes. Our findings should greatly advance our understanding of the adaptive mechanisms responsible for maintaining cell integrity in times of sustained environmental shocks.


Assuntos
Actinobacteria , Actinomycetales , Muramidase/metabolismo , Peptidoglicano/metabolismo , Etambutol/metabolismo , Isoniazida/metabolismo , Actinomycetales/genética , Parede Celular/metabolismo , Actinobacteria/genética , Actinobacteria/metabolismo , Antibacterianos/farmacologia , Antibacterianos/metabolismo , beta-Lactamas/metabolismo , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Regulação Bacteriana da Expressão Gênica
4.
Biotechnol Lett ; 44(12): 1477-1493, 2022 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-36315299

RESUMO

OBJECTIVES: As a type of agricultural waste, there is a large amount of lignocellulose in corn (Zea mays) straw, but it is difficult to utilize efficiently owing to its recalcitrance to enzymatic degradation. Three strains of actinomycetes that degrade cellulose were constructed as complex flora, and the conditions of cellulose degradation conditions and their degradative activity were optimized and evaluated. RESULTS: When the complex flora were inoculated into the fermentation medium at pH 7 and 3% (v/v), the rate of degradation of corn straw reached 38.24% after 5 d of fermentation at 28 ºC and 180 rpm. Cellulose, hemicellulose, and lignin in the corn straw were degraded by 33.97%, 34.08%, and 21.52%, respectively. The results from scanning electron microscopy showed that the waxy layer on the surface of corn straw became thin and gradually disappeared following fermentation by the complex flora. Fourier-transform infrared spectroscopy showed that the complex flora could change the internal functional groups of corn straw at different fermentation periods. The compounds detected in the fermentation system indicated that the corn straw was efficiently degraded. CONCLUSIONS: These results indicated that the constructed complex flora was more effective at degrading corn straw than the individual strains and provides research concepts for the development and utilization of biomass resources.


Assuntos
Actinobacteria , Zea mays , Zea mays/química , Actinobacteria/metabolismo , Actinomyces , Celulose/metabolismo , Fermentação
5.
World J Microbiol Biotechnol ; 38(12): 250, 2022 Oct 29.
Artigo em Inglês | MEDLINE | ID: mdl-36308608

RESUMO

Actinobacteria are well-known Gram-positive bacteria that produce approximately two-thirds of microbial bioactive natural products (NPs) through secondary metabolism. Usually, genes involved in the biosynthesis of NPs in actinobacteria are clustered, and their expression is regulated by an elaborate and stringent regulatory network formed by diverse regulators. These regulators can be classified into more than 50 superfamilies/families according to conserved amino acid sequences and biological functions. Among them, LuxR family regulators, which are widely distributed in microorganisms and feature an HTH_LUXR domain (PFAM00196, SMART00421), play key roles in quorum sensing (QS), bioluminescence, virulence and secondary metabolism. In this mini-review, we focus on their roles in regulating NP production in actinobacteria. First, the domain architecture and classification of LuxR proteins are summarized on the basis of their size and biological function diversity. Second, the landscape of the roles and action mechanism of LuxR regulators involved in NP production in actinobacteria is presented in detail. Finally, the application of LuxR is described from two perspectives: enhancement of NP production and discovery of novel NPs by engineering LuxR. This mini-review will help us comprehensively understand the role of LuxR in actinobacteria and promote the future application of LuxR family regulators in synthetic biology.


Assuntos
Actinobacteria , Humanos , Actinobacteria/genética , Actinobacteria/metabolismo , Transativadores/genética , Transativadores/metabolismo , Proteínas Repressoras/genética , Proteínas Repressoras/metabolismo , Percepção de Quorum/genética , Bactérias/metabolismo , Regulação Bacteriana da Expressão Gênica , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo
6.
Bioresour Technol ; 363: 127949, 2022 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-36108576

RESUMO

This study was the first to explore the effect of shell powder (SP) on lignin degradation and humus (HS) formation during composting. The results showed that the treatment group (T) with SP consumed more polyphenols, reducing sugar and amino acids than the control group (CK), especially the rate of reducing sugar consumption in T (50.61 %) was significantly higher than CK (28.40 %). SP greatly enhanced the efficiency of lignin degradation (T:45.47 %; CK:24.63 %) and HS formation (T:34.93 %; CK:20.16 %). The content of HA in T was 12.94 mg/g while CK was 12.06 mg/g. SP maintained a continuous increase in the relative abundance of AA1, AA3 after cooling phase. Meanwhile, T (48.98 %) significantly increased the abundance of Actinobacteria compared with CK (37.19 %). Actinobacteria, AA1 and AA3 were identified as the main factors promoting lignin degradation and HS formation by correlation analysis. Therefore, adding SP could be a novel strategy to improve compost quality.


Assuntos
Actinobacteria , Compostagem , Actinobacteria/metabolismo , Aminoácidos , Bactérias/metabolismo , Açúcares da Dieta , Lignina/metabolismo , Esterco , Pós , Solo , Açúcares
7.
J Environ Manage ; 323: 116220, 2022 Dec 01.
Artigo em Inglês | MEDLINE | ID: mdl-36116255

RESUMO

Rhodococcus is a genus of actinomycetes that has been explored by the scientific community for different purposes, especially for bioremediation uses. However, the mechanisms governing Rhodococcus-mediated bioremediation processes are far from being fully elucidated. In this sense, this work aimed to compile the recent advances in the use of Rhodococcus for the bioremediation of organic and inorganic contaminants present in different environmental compartments. We reviewed the bioremediation capacity and mechanisms of Rhodococcus spp. in the treatment of polycyclic aromatic hydrocarbons, phenolic substances, emerging contaminants, heavy metals, and dyes given their human health risks and environmental concern. Different bioremediation techniques were discussed, including experimental conditions, treatment efficiencies, mechanisms, and degradation pathways. The use of Rhodococcus strains in the bioremediation of several compounds is a promising approach due to their features, primarily the presence of appropriate enzyme systems, which result in high decontamination efficiencies; but that vary according to experimental conditions. Besides, the genus Rhodococcus contains a small number of opportunistic species and pathogens, representing an advantage from the point of view of safety. Advances in analytical detection techniques and Molecular Biology have been collaborating to improve the understanding of the mechanisms and pathways involved in bioremediation processes. In the context of using Rhodococcus spp. as bioremediation agents, there is a need for more studies that 1) evaluate the role of these actinomycetes on a pilot and field scale; 2) use genetic engineering tools and consortia with other microorganisms to improve the bioremediation efficiency; and 3) isolate new Rhodococcus strains from environments with extreme and/or contaminated conditions aiming to explore their adaptive capabilities for bioremediation purposes.


Assuntos
Actinobacteria , Metais Pesados , Hidrocarbonetos Policíclicos Aromáticos , Rhodococcus , Actinobacteria/metabolismo , Actinomyces/metabolismo , Biodegradação Ambiental , Corantes/metabolismo , Humanos , Metais Pesados/metabolismo , Hidrocarbonetos Policíclicos Aromáticos/metabolismo , Rhodococcus/genética , Rhodococcus/metabolismo
8.
Int J Mol Sci ; 23(18)2022 Sep 19.
Artigo em Inglês | MEDLINE | ID: mdl-36142891

RESUMO

The bile resistance of intestinal bacteria is among the key factors responsible for their successful colonization of and survival in the mammalian gastrointestinal tract. In this study, we demonstrated that lactate-producing Atopobiaceae bacteria (Leptogranulimonas caecicola TOC12T and Granulimonas faecalis OPF53T) isolated from mouse intestine showed high resistance to mammalian bile extracts, due to significant bile salt hydrolase (BSH) activity. We further succeeded in isolating BSH proteins (designated LcBSH and GfBSH) from L. caecicola TOC12T and G. faecalis OPF53T, respectively, and characterized their enzymatic features. Interestingly, recombinant LcBSH and GfBSH proteins exhibited BSH activity against 12 conjugated bile salts, indicating that LcBSH and GfBSH have much broader substrate specificity than the previously identified BSHs from lactic acid bacteria, which are generally known to hydrolyze six bile salt isomers. Phylogenetic analysis showed that LcBSH and GfBSH had no affinities with any known BSH subgroup and constituted a new BSH subgroup in the phylogeny. In summary, we discovered functional BSHs with broad substrate specificity from Atopobiaceae bacteria and demonstrated that these BSH enzymes confer bile resistance to L. caecicola TOC12T and G. faecalis OPF53T.


Assuntos
Actinobacteria , Lactobacillales , Actinobacteria/metabolismo , Amidoidrolases/metabolismo , Animais , Bile/metabolismo , Ácidos e Sais Biliares , Lactatos , Lactobacillales/metabolismo , Mamíferos/metabolismo , Camundongos , Filogenia , Especificidade por Substrato
9.
J Environ Manage ; 320: 115870, 2022 Oct 15.
Artigo em Inglês | MEDLINE | ID: mdl-36056489

RESUMO

Biopurification systems (BPS) or biobeds are bioprophylaxis systems to prevent pesticide point-source contamination, whose efficiency relies mostly on the pesticide removal capacity of the biomixture, the majority component of a BPS. The adaptation of the components of the biomixtures to local availabilities is a key aspect to ensure the sustainability of the system. In this work, the removal of atrazine (ATZ) was evaluated in biomixtures formulated with three sugarcane by-products as alternative lignocellulosic substrates. Based on the capacity of actinobacteria to tolerate and degrade diverse pesticides, the effect of biomixtures bioaugmentation with actinobacteria was evaluated as a strategy to enhance the depuration capacity of biobeds. Also, the effect of ATZ and/or the bioaugmentation on microbial developments and enzymatic activities were studied. The biomixtures formulated with bagasse, filter cake, or harvest residue, reached pesticide removal values of 37-41% at 28 d of incubation, with t1/2 between 37.9 ± 0.4 d and 52.3 ± 0.4 d. The bioaugmentation with Streptomyces sp. M7 accelerated the dissipation of the pesticide in the biomixtures, reducing ATZ t1/2 3-fold regarding the controls, and achieving up to 72% of ATZ removal. Atrazine did not exert a clear effect on microbial developments, although most of the microbial counts were less in the contaminated biomixtures at the end of the assay. The bioaugmentation improved the development of the microbiota in general, specially actinobacteria and fungi, regarding the non-bioaugmented systems. The inoculation with Streptomyces sp. M7 enhanced acid phosphatase activity and/or reversed a possible effect of the pesticide over this enzymatic activity.


Assuntos
Actinobacteria , Atrazina , Praguicidas , Poluentes do Solo , Streptomyces , Actinobacteria/metabolismo , Atrazina/metabolismo , Biodegradação Ambiental , Solo/química , Poluentes do Solo/metabolismo , Streptomyces/metabolismo
10.
Environ Res ; 214(Pt 3): 114018, 2022 11.
Artigo em Inglês | MEDLINE | ID: mdl-35961544

RESUMO

Every year, 180 billion tonnes of cellulose are produced by plants as waste biomass after the cultivation of the desired product. One of the smart and effective ways to utilize this biomass rather than burn it is to utilize the biomass to adequately meet the energy needs with the help of microbial cellulase that can catalytically convert the cellulose into simple sugar units. Marine actinobacteria is one of the plentiful gram-positive bacteria known for its industrial application as it can produce multienzyme cellulase with high thermal tolerance, pH stability and high resistant towards metal ions and salt concentration, along with other antimicrobial properties. Highly stable cellulase obtained from marine actinobacteria will convert the cellulose biomass into glucose, which is the precursor for biofuel production. This review will provide a comprehensive outlook of various strategic applications of cellulase from marine actinobacteria which can facilitate the breakdown of lignocellulosic biomass to bioenergy with respect to its characteristics based on the location/environment that the organism was collected and its screening strategies followed by adopted methodologies to mine the novel cellulase genome and enhance the production, thereby increasing the activity of cellulase continued by effective immobilization on novel substrates for the multiple usage of cellulase along with the industrial applications.


Assuntos
Actinobacteria , Celulase , Actinobacteria/genética , Actinobacteria/metabolismo , Bactérias/metabolismo , Biocombustíveis , Biomassa , Celulase/química , Celulase/genética , Celulase/metabolismo , Celulose/metabolismo
11.
J Appl Microbiol ; 133(4): 2417-2429, 2022 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-35808848

RESUMO

AIMS: Actinobacteria are known to produce extracellular enzymes including DyPs. We set out to identify and characterize novel peroxidases from Streptomyces chartreusis NRRL 3882, because S. chartreusis belongs to the small group of actinobacteria with three different DyPs. METHODS AND RESULTS: The genome of the actinomycete S. chartreusis NRRL 3882 was mined for novel DyP-type peroxidases. Three genes encoding for DyP-type peroxidases were cloned and overexpressed in Escherichia coli. Subsequent characterization of the recombinant proteins included examination of operating conditions such as pH, temperature and H2 O2 concentrations, as well as substrate spectrum. Despite their high sequence similarity, the enzymes named SCDYP1-SCDYP3 presented distinct preferences regarding their operating conditions. They showed great divergence in H2 O2 tolerance and stability, with SCDYP2 being most active at concentrations above 50 mmol l-1 . Moreover, SCDYP1 and SCDYP3 preferred acidic pH (typical for DyP-type peroxidases), whereas SCDYP2 was most active at pH 8. CONCLUSIONS: Regarding the function of DyPs in nature, these results suggest that availability of different DyP variants with complementary activity profiles in one organism might convey evolutionary benefits. SIGNIFICANCE AND IMPACT OF THE STUDY: DyP-type peroxidases are able to degrade xenobiotic compounds and thus can be applied in biocatalysis and bioremediation. However, the native function of DyPs and the benefits for their producers largely remain to be elucidated.


Assuntos
Actinobacteria , Peroxidases , Actinobacteria/genética , Actinobacteria/metabolismo , Corantes/metabolismo , Escherichia coli/genética , Escherichia coli/metabolismo , Peroxidases/genética , Peroxidases/metabolismo , Proteínas Recombinantes/metabolismo , Streptomyces , Xenobióticos/metabolismo
12.
Environ Sci Pollut Res Int ; 29(58): 88331-88346, 2022 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-35834084

RESUMO

Indigenous microbial diversity has potential for rapid decomposition of residue through enzyme activities that is alternative, effective, and environment friendly strategy to accelerate degradation of lignocellulose in agricultural residues and make composting process economically viable. Keeping this view, the main objective of the present study was isolation and characterization of lignocellulosic degrading microbial diversity from long-term residue management practice experiments and to develop potential microbial consortium for rapid degradation of lignocellulosic biomass. In this study, twenty-five bacteria, nine fungi, and four actinomycetes isolates were obtained from the soil samples of different residue management fields from Ludhiana, Punjab, India. All isolates were qualitatively and quantitatively screened for enzyme activities, i.e., cellulase, xylanase, laccase, and lignin peroxidase. On the basis of quantitative estimation of enzyme activities, 3 fungal (S1F1, S2F4, and S6F9), 2 actinomycetes (S1A1 and S6A4), and 2 bacterial strains (S6B16 and S6B17) were further selected for in vitro bio-compatibility assay. Selected bio-compatible microbial strains were identified as Streptomyces flavomacrosporus (S6A4), Aspergillus terreus (S2F4), and Bacillus altitudinis (S6B16) through 16S rRNA and 18S rRNA sequencing. Furthermore, single and developed microbial consortium (S6B16 + S6A4 + S2F4) were screened for quantitative estimation of cellulase, xylanase, laccase, and lignin peroxidase enzymes with 23 biochemically different cereal, legume, and oil seed crop residues for optimization of enzyme activities at different time intervals. Results revealed that Vigna radiata followed by Cajanus cajan and Arachis hypogaea straw residue powder @ 1% in culture broth are a promising carbon source for B. altitudinis, S. flavomacrosporus, and A. terreus to produce higher ligno-cellulolytic microbial degrading enzymes due to variable range of carbon (C):nitrogen (N) ratio and higher ligno-cellulolytic content in the studied crop residues. Thus, the application of indigenous microbial consortium with efficient lignocellulose hydrolysis enzyme machinery might be an attractive alternative for ex situ crop residue management practices under sustainable manners.


Assuntos
Actinobacteria , Celulase , Consórcios Microbianos , RNA Ribossômico 16S , Lacase , Lignina/metabolismo , Celulase/metabolismo , Actinobacteria/metabolismo , Carbono
13.
Proc Natl Acad Sci U S A ; 119(28): e2203114119, 2022 07 12.
Artigo em Inglês | MEDLINE | ID: mdl-35787040

RESUMO

Most Actinobacteria encode a small transmembrane protein, whose gene lies immediately downstream of the housekeeping sortase coding for a transpeptidase that anchors many extracellular proteins to the Gram-positive bacterial cell wall. Here, we uncover the hitherto unknown function of this class of conserved proteins, which we name SafA, as a topological modulator of sortase in the oral Actinobacterium Actinomyces oris. Genetic deletion of safA induces cleavage and excretion of the otherwise predominantly membrane-bound SrtA in wild-type cells. Strikingly, the safA mutant, although viable, exhibits severe abnormalities in cell morphology, pilus assembly, surface protein localization, and polymicrobial interactions-the phenotypes that are mirrored by srtA depletion. The pleiotropic defect of the safA mutant is rescued by ectopic expression of safA from not only A. oris, but also Corynebacterium diphtheriae or Corynebacterium matruchotii. Importantly, the SrtA N terminus harbors a tripartite-domain feature typical of a bacterial signal peptide, including a cleavage motif AXA, mutations in which prevent SrtA cleavage mediated by the signal peptidase LepB2. Bacterial two-hybrid analysis demonstrates that SafA and SrtA directly interact. This interaction involves a conserved motif FPW within the exoplasmic face of SafA, since mutations of this motif abrogate SafA-SrtA interaction and induce SrtA cleavage and excretion as observed in the safA mutant. Evidently, SafA is a membrane-imbedded antagonist of signal peptidase that safeguards and maintains membrane homeostasis of the housekeeping sortase SrtA, a central player of cell surface assembly.


Assuntos
Actinobacteria/metabolismo , Aminoaciltransferases , Aminoaciltransferases/genética , Aminoaciltransferases/metabolismo , Proteínas de Bactérias/metabolismo , Cisteína Endopeptidases/genética , Cisteína Endopeptidases/metabolismo , Homeostase , Proteínas de Membrana , Morfogênese , Serina Endopeptidases
14.
Environ Sci Pollut Res Int ; 29(50): 76298-76309, 2022 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-35668255

RESUMO

The application of ionic liquids with sawdust and fresh dairy manure was studied in composting. The degradation of organic matter (OM), dissolved organic matter (DOM), and lignocellulose was analyzed. The DOM decreased by 14.25 mg/g and 11.11 mg/g in experimental group (ILs) and control group (CK), respectively. OM decreased by 7.32% (CK) and 8.91% (ILs), respectively. The degradation rates of hemicellulose, lignin, and cellulose in ILs (56.62%, 42.01%, and 23.97%) were higher than in CK (38.39%, 39.82%, and 16.04%). Microbial community and carbohydrate-active enzymes (CAZymes) were analyzed based on metagenomics. Metagenomic analysis results showed that ionic liquids enriched Actinobacteria and Proteobacteria in composting. Compared with CK, the total abundance values of GH11, GH6, AA6, and AA3_2 in ILs increased by 13.98%, 10.12%, 11.21%, and 13.68%, respectively. Ionic liquids can improve the lignocellulosic degradation by regulating the environmental physicochemical parameters (temperature, pH, C/N) to promote the growth of Actinobacteria and Proteobacteria and carbohydrate-active enzymes (CAZymes) abundance. Therefore, ionic liquids are a promising additive in lignocellulosic waste composting.


Assuntos
Actinobacteria , Compostagem , Líquidos Iônicos , Microbiota , Actinobacteria/genética , Actinobacteria/metabolismo , Bactérias/genética , Bactérias/metabolismo , Celulose , Lignina/metabolismo , Esterco/microbiologia , Metagenômica , Solo
15.
ACS Synth Biol ; 11(6): 2022-2028, 2022 06 17.
Artigo em Inglês | MEDLINE | ID: mdl-35674818

RESUMO

Biochemical and structural analyses of purified proteins are essential for the understanding of their properties. However, many proteins are unstable and difficult to purify, hindering their characterization. The B2 proteins of the lasso peptide biosynthetic pathways are cysteine proteases that cleave precursor peptides during the maturation process. The B2 proteins are poorly soluble, and no experimentally solved structures are available. Here, we performed a rapid semicomprehensive mutational analysis of the B2 protein from the thermophilic actinobacterium, Thermobifida fusca (FusB2), using a cell-free transcription/translation system, and compared the results with the structure prediction by AlphaFold2. Analysis of 34 FusB2 mutants with substitutions of hydrophobic residues confirmed the accuracy of the predicted structure, and revealed a hydrophobic patch on the protein surface, which likely serves as the binding site of the partner protein, FusB1. Our results suggest that the combination of rapid cell-free mutant analyses with precise structure predictions can greatly accelerate structure-function research of proteins for which no structures are available.


Assuntos
Actinobacteria , Peptídeo Hidrolases , Actinobacteria/metabolismo , Endopeptidases , Peptídeos/metabolismo , Proteínas
16.
Angew Chem Int Ed Engl ; 61(32): e202205409, 2022 08 08.
Artigo em Inglês | MEDLINE | ID: mdl-35656913

RESUMO

Benzoxazole scaffolds feature prominently in diverse synthetic and natural product-derived pharmaceuticals. Our understanding of their bacterial biosynthesis is, however, limited to ortho-substituted heterocycles from actinomycetes. We report an overlooked biosynthetic pathway in anaerobic bacteria (typified in Clostridium cavendishii) that expands the benzoxazole chemical space to meta-substituted heterocycles and heralds a distribution beyond Actinobacteria. The first benzoxazoles from the anaerobic realm (closoxazole A and B) were elucidated by NMR and chemical synthesis. By genome editing in the native producer, heterologous expression in Escherichia coli, and systematic pathway dissection we show that closoxazole biosynthesis invokes an unprecedented precursor usage (3-amino-4-hydroxybenzoate) and manner of assembly. Synthetic utility was demonstrated by the precursor-directed biosynthesis of a tafamidis analogue. A bioinformatic survey reveals the pervasiveness of related gene clusters in diverse bacterial phyla.


Assuntos
Actinobacteria , Bactérias Anaeróbias , Actinobacteria/metabolismo , Bactérias/metabolismo , Bactérias Anaeróbias/genética , Benzoxazóis/química , Vias Biossintéticas/genética , Escherichia coli/metabolismo , Família Multigênica
17.
Sci Rep ; 12(1): 7222, 2022 05 04.
Artigo em Inglês | MEDLINE | ID: mdl-35508597

RESUMO

Co-culture is an efficient strategy for natural product discovery. We have used mycolic acid-containing bacteria (MACB) Tsukamurella pumonis TP-B0596 to induce secondary metabolism by actinomycetes and have found several natural products. We also observed that MACB attached to the mycelium of Streptomyces lividans forming coaggregates during combined-culture. This stimulated interest in the interactions among actinomycetes and MACB, and we found that soil isolated cultures contained a mixture of actinomycetes and MACB. Our previously observed interactions were the result of selective screening and combination of bacteria in the lab, which warranted investigation of the existence of these interactions in the natural soil environment. Therefore, in this paper, we report the interaction between a co-isolated natural pair of actinomycetes and MACB in terms of morphology and metabolic changes. A natural pair of actinomycetes and MACB co-aggregated in liquid culture and showed metabolic changes. Interestingly, co-aggregated actinomycetes and MACB were re-isolated from soil with no obvious morphological colony differences from the colony of a single strain. The results demonstrate that there is a stochastic chance of picking colonies containing co-aggregated actinomycetes and MACB, which suggests that the pair can exist in co-aggregate form in the soil environment and interact with each other.


Assuntos
Actinobacteria , Produtos Biológicos , Actinobacteria/metabolismo , Actinomyces/metabolismo , Bactérias/metabolismo , Produtos Biológicos/metabolismo , Ácidos Micólicos/metabolismo , Solo
18.
mBio ; 13(3): e0039322, 2022 06 28.
Artigo em Inglês | MEDLINE | ID: mdl-35608300

RESUMO

Bacterial natural products have historically been a deep source of new medicines, but their slowed discovery in recent decades has put a premium on developing strategies that enhance the likelihood of capturing novel compounds. Here, we used a straightforward approach that capitalizes on the interactive ecology of "rare" actinomycetes. Specifically, we screened for interactions that triggered the production of antimicrobials that inhibited the growth of a bacterial strain with exceptionally diverse natural antimicrobial resistance. This strategy led to the discovery of a family of antimicrobials we term the dynaplanins. Heterologous expression enabled identification of the dynaplanin biosynthetic gene cluster, which was missed by typical algorithms for natural product gene cluster detection. Genome sequencing of partially resistant mutants revealed a 2-oxo acid dehydrogenase E2 subunit as the likely molecular target of the dynaplanins, and this finding was supported by computational modeling of the dynaplanin scaffold within the active site of this enzyme. Thus, this simple strategy, which leverages microbial interactions and natural antibiotic resistance, can enable discovery of molecules with unique antimicrobial activity. In addition, these results indicate that primary metabolism may be a direct target for inhibition via chemical interference in competitive microbial interactions. IMPORTANCE Many antibiotics were originally discovered from microbes. However, in recent decades, resistance to current treatments has risen, while novel antibiotic discovery has become increasingly challenging. Thus, there is a need to develop new strategies to find novel antimicrobials. Here, we incorporated three levels of innovation into a single, simple discovery pipeline: focusing on understudied bacteria with a high potential for producing antibiotics, growing these bacteria in binary microbial interactions, and screening for activity against a multidrug-resistant bacterium. This led us to discover a family of antimicrobials that we call the dynaplanins, which are synthesized by genes that were not detected by typical prediction algorithms. We found that dynaplanins likely block the function of one of three related enzymes called 2-oxo acid dehydrogenases, which are vital to cellular metabolism. Overall, our strategy based on bacterial competition led to discovery of a novel antibiotic that inhibits the ability to metabolize nutrients.


Assuntos
Actinobacteria , Produtos Biológicos , Actinobacteria/genética , Actinobacteria/metabolismo , Antibacterianos/metabolismo , Bactérias/metabolismo , Produtos Biológicos/metabolismo , Produtos Biológicos/farmacologia , Farmacorresistência Bacteriana , Cetoácidos
19.
Int J Mol Sci ; 23(10)2022 May 18.
Artigo em Inglês | MEDLINE | ID: mdl-35628478

RESUMO

The human society faces a serious problem due to the widespread resistance to antibiotics in clinical practice. Most antibiotic biosynthesis gene clusters in actinobacteria contain genes for intrinsic self-resistance to the produced antibiotics, and it has been proposed that the antibiotic resistance genes in pathogenic bacteria originated in antibiotic-producing microorganisms. The model actinobacteria Streptomyces clavuligerus produces the ß-lactam antibiotic cephamycin C, a class A ß-lactamase, and the ß lactamases inhibitor clavulanic acid, all of which are encoded in a gene supercluster; in addition, it synthesizes the ß-lactamase inhibitory protein BLIP. The secreted clavulanic acid has a synergistic effect with the cephamycin produced by the same strain in the fight against competing microorganisms in its natural habitat. High levels of resistance to cephamycin/cephalosporin in actinobacteria are due to the presence (in their ß-lactam clusters) of genes encoding PBPs which bind penicillins but not cephalosporins. We have revised the previously reported cephamycin C and clavulanic acid gene clusters and, in addition, we have searched for novel ß-lactam gene clusters in protein databases. Notably, in S. clavuligerus and Nocardia lactamdurans, the ß-lactamases are retained in the cell wall and do not affect the intracellular formation of isopenicillin N/penicillin N. The activity of the ß-lactamase in S. clavuligerus may be modulated by the ß-lactamase inhibitory protein BLIP at the cell-wall level. Analysis of the ß-lactam cluster in actinobacteria suggests that these clusters have been moved by horizontal gene transfer between different actinobacteria and have culminated in S. clavuligerus with the organization of an elaborated set of genes designed for fine tuning of antibiotic resistance and cell wall remodeling for the survival of this Streptomyces species. This article is focused specifically on the enigmatic connection between ß-lactam biosynthesis and ß-lactam resistance mechanisms in the producer actinobacteria.


Assuntos
Actinobacteria , Proteínas de Ligação às Penicilinas , Inibidores de beta-Lactamases , beta-Lactamases , Actinobacteria/genética , Actinobacteria/metabolismo , Antibacterianos/farmacologia , Cefalosporinas/farmacologia , Cefamicinas/farmacologia , Ácido Clavulânico/farmacologia , Proteínas de Ligação às Penicilinas/metabolismo , Penicilinas/farmacologia , Inibidores de beta-Lactamases/farmacologia , beta-Lactamases/metabolismo , beta-Lactamas/farmacologia
20.
Methods Mol Biol ; 2489: 157-171, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-35524050

RESUMO

Bacteria produce an impressive array of bioactive specialized metabolites, with Streptomyces (and the actinobacteria more generally) being unusually diverse and prolific producers. However, the biosynthetic potential of these organisms has yet to be fully explored, as many of the biosynthetic gene clusters that direct the synthesis of these natural products are transcriptionally silent under laboratory growth conditions. Here, we describe strategies that can be employed to broadly stimulate the expression of biosynthetic gene clusters in Streptomyces and their relatives, follow the transcription of these genes, and assess the antimicrobial activity of the resulting molecules.


Assuntos
Actinobacteria , Produtos Biológicos , Streptomyces , Actinobacteria/genética , Actinobacteria/metabolismo , Produtos Biológicos/metabolismo , Família Multigênica , Streptomyces/genética , Streptomyces/metabolismo
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