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1.
Appl Microbiol Biotechnol ; 105(18): 6759-6778, 2021 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-34458936

RESUMO

The genus Cohnella belongs to a group of Gram-positive endospore-forming bacteria within the Paenibacillaceae family. Although most species were described as xylanolytic bacteria, the literature still lacks some key information regarding their repertoire of xylan-degrading enzymes. The whole genome sequence of an isolated xylan-degrading bacterium Cohnella sp. strain AR92 was found to contain five genes encoding putative endo-1,4-ß-xylanases, of which four were cloned, expressed, and characterized to better understand the contribution of the individual endo-xylanases to the overall xylanolytic properties of strain AR92. Three of the enzymes, CoXyn10A, CoXyn10C, and CoXyn11A, were shown to be effective at hydrolyzing xylans-derived from agro-industrial, producing oligosaccharides with substrate conversion values of 32.5%, 24.7%, and 10.6%, respectively, using sugarcane bagasse glucuronoarabinoxylan and of 29.9%, 19.1%, and 8.0%, respectively, using wheat bran-derived arabinoxylan. The main reaction products from GH10 enzymes were xylobiose and xylotriose, whereas CoXyn11A produced mostly xylooligosaccharides (XOS) with 2 to 5 units of xylose, often substituted, resulting in potentially prebiotic arabinoxylooligosaccharides (AXOS). The endo-xylanases assay displayed operational features (temperature optima from 49.9 to 50.4 °C and pH optima from 6.01 to 6.31) fitting simultaneous xylan utilization. Homology modeling confirmed the typical folds of the GH10 and GH11 enzymes, substrate docking studies allowed the prediction of subsites (- 2 to + 1 in GH10 and - 3 to + 1 in GH11) and identification of residues involved in ligand interactions, supporting the experimental data. Overall, the Cohnella sp. AR92 endo-xylanases presented significant potential for enzymatic conversion of agro-industrial by-products into high-value products.Key points• Cohnella sp. AR92 genome encoded five potential endo-xylanases.• Cohnella sp. AR92 enzymes produced xylooligosaccharides from xylan, with high yields.• GH10 enzymes from Cohnella sp. AR92 are responsible for the production of X2 and X3 oligosaccharides.• GH11 from Cohnella sp. AR92 contributes to the overall xylan degradation by producing substituted oligosaccharides.


Assuntos
Bacillales , Saccharum , Endo-1,4-beta-Xilanases/genética , Hidrólise , Oligossacarídeos , Xilanos
2.
Arch Microbiol ; 203(8): 4859-4865, 2021 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-34235583

RESUMO

A Gram-positive, rod-shaped, spore-forming, thermophilic, and acidophilic bacterium, designated as strain skT53T, was isolated from farm soil in Tokyo, Japan. Under aerobic conditions, the strain grew at 35-55 °C (optimum temperature 44-55 °C) and pH 4.0-6.0 (optimum pH 5.0). Phylogenetic analysis of the 16S rRNA gene sequence showed that the isolate was moderately related to the type strain of Effusibacillus consociatus (94.3% similarity). The G + C content of the genomic DNA was 48.2 mol%, and MK-7 was the predominant respiratory quinone in the strain. The major fatty acids were anteiso-C15:0, iso-C15:0, and iso-C16:0. Based on the phenotypic and chemotaxonomic characteristics, as well as 16S rRNA gene sequence similarity and whole genome analyses, strain skT53T represents a novel species in the genus Effusibacillus, for which the name Effusibacillus dendaii sp. nov. has been proposed. The type strain is skT53T (= NBRC 114101 T = TBRC 11241 T).


Assuntos
Microbiologia do Solo , Solo , Bacillales , Técnicas de Tipagem Bacteriana , Composição de Bases , DNA Bacteriano/genética , Fazendas , Ácidos Graxos , Filogenia , RNA Ribossômico 16S/genética , Análise de Sequência de DNA
3.
Enzyme Microb Technol ; 149: 109854, 2021 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-34311891

RESUMO

Xylanases are categorized into different family groups, two of which are glycoside hydrolases 10 (GH10) and 11 (GH11) families. These well-characterized xylanases demonstrate different modes of action in hydrolysis of xylans. Imitating certain types of microorganisms to produce bifunctional enzymes such as engineered xylanases has gained considerable attention among researchers. In this study, a recombinant chimeric enzyme (X11-10) was designed by fusing two thermostable xylanases through a peptide linker. The recombinant parental enzymes, xylanase 10 from fungus Bispora sp. MEY-1 (X10) and xylanase 11 from bacterium Thermobacillus xylanilyticus (X11), and their chimera were successfully expressed in Pichia pastoris (P. pastoris), purified, and characterized. Being active over a wide pH range, X11-10 chimera showed higher thermal stability, possessed a lower Km, and a higher catalytic efficiency (kcat/Km) in comparison to the parental enzymes. Also, molecular dynamics simulation (MDS) of X11-10 revealed that its active site residues were free to interact with substrate. This novel chimeric xylanase may have potential applications in different industrial processes since it can substitute two separate enzymes and therefore minimize the production costs.


Assuntos
Ascomicetos , Xilanos , Ascomicetos/metabolismo , Bacillales , Quimera/metabolismo , Endo-1,4-beta-Xilanases/genética , Endo-1,4-beta-Xilanases/metabolismo , Estabilidade Enzimática , Glicosídeo Hidrolases , Proteínas Recombinantes de Fusão/genética , Saccharomycetales , Especificidade por Substrato
4.
J Environ Manage ; 294: 113162, 2021 Sep 15.
Artigo em Inglês | MEDLINE | ID: mdl-34214942

RESUMO

The plant residues of tomato bring pressures to the environment and composting provides a feasible method to treat such agricultural waste. However, little is known about the succession and associations of the dominant lignocellulose degraders in the compost system. To further accelerate the process by inoculating key functional microorganisms, a compost pile composed of tomato stalk with maize straw addition was constructed, and the whole community structure and functions of the dominant were investigated by applying the integrated mata-omics. Results showed that Actinobacteria, Firmicutes, and Ascomycota dominated and drove the assembly of the co-occurrence network. In the thermophilic stage, Thermobifida was the exclusive degrader of cellulose, and Thermobifida fusca was the most important cellulolytic actinomycete. Saccharomonospora viridis, Planifilum fulgidum, Thermobacillus sp. and the dominant ascomycota of Aspergillus sclerotialis participated in hemicellulose decomposing. In the cooling phase, functional microorganisms became more diverse, with Nocardiopsis flavescens, Glycomyces artemisiae, Glycomyces sambucus, Streptomyces rubrolavendulae and Streptomyces vietnamensis joining the cellulose-degrading rank, and Chaetomium thermophilum emerging as the main hemicellulose degrader. More than two thirds of the bacteria-bacteria interactions and all the fungi-fungi associations were positive, while, both competition (for the same substrate of hemicellulose) and synergy (preference for cellulose and hemicellulose) coexisted in the bacteria-fungi interactions. In conclusion, these findings provide useful information for understanding the biodegradation of tomato plant residues better, and effects of the functional agents identified on composting process should be further studied.


Assuntos
Compostagem , Lycopersicon esculentum , Actinobacteria , Aspergillus , Bacillales , Chaetomium , Lignina , Solo , Streptomyces
6.
Artigo em Inglês | MEDLINE | ID: mdl-34161218

RESUMO

A Gram-positive, aerobic, flagellated, endospore-forming, rod-shaped strain, designated as G13T, was isolated from soil. The results of 16S rRNA gene sequence analysis led to the conclusion that strain G13T was phylogenetically related to Cohnella boryungensis BR29T (97.5 %) and Cohnella phaseoli CECT 7287T (96.9 %) with digital DNA-DNA hybridization values of 21.0 and 21.4 %, and distantly related to Cohnella thermotolerans CCUG 47242T (94.8 %), type species of the genus Cohnella, at 19.0 %. The genome size of strain G13T was 5 387 258 bp, with 51.3 mol% G+C content. The predominant fatty acids were summed feature 9 (iso-C17 : 1 ω9c and/or C16 : 0 10-methyl), anteiso-C17 : 0, iso-C17 : 0 and iso-C15 : 0. The predominant quinone was menaquinone-7 and the major polar lipids were diphosphatidyglycerol, phosphatidylethanolamine, phosphatidylglycerol, lysylphosphatidylglycerol, three aminophospholipids, two phosphoglycolipids, three aminolipids and two unidentified lipids. Based on the data from phenotypic tests and the genotypic differences between strain G13T and its close phylogenetic relatives, strain G13T represents a new species belonging to the genus Cohnella, for which the name Cohnella terricola sp. nov. (=KACC 19905T=NBRC 113748T) is proposed.


Assuntos
Bacillales/classificação , Filogenia , Microbiologia do Solo , Bacillales/isolamento & purificação , Técnicas de Tipagem Bacteriana , Composição de Bases , DNA Bacteriano/genética , Ácidos Graxos/química , Hibridização de Ácido Nucleico , Fosfolipídeos/química , RNA Ribossômico 16S/genética , República da Coreia , Análise de Sequência de DNA , Vitamina K 2/análogos & derivados , Vitamina K 2/química
7.
Arch Microbiol ; 203(6): 3577-3590, 2021 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-33961074

RESUMO

Recently, the industrial-scale development of microbial D-lactic acid production has been discussed. In this study, the efficiency of the new isolate Sporolactobacillus terrae SBT-1 for producing D-lactic acid under challenge conditions was investigated. The isolate SBT-1 exhibited superior activity in fermenting a very high glucose or sucrose concentration to D-lactic acid compared to the other S. terrae isolates previously reported in the literature; therefore, SBT-1 could overcome the limitations of effective lactic acid production. In batch cultivation using 360 g/L glucose, SBT-1 produced 290.30 g/L D-lactate with a sufficiently high glucose conversion yield of 86%, volumetric productivity of 3.02 g/L h, and optical purity of 96.80% enantiomer excess. SBT-1 could also effectively utilize 440 g/L sucrose as a sole carbon source to produce 276.50 g/L lactic acid with a conversion yield of 90%, a production rate of 2.88 g/L h, and an optical purity of 98%. D-Lactic acid fermentation by two other related producers, S. inulinus NRIC1133T and S. terrae NRIC0357T, was compared with fermentation by isolate SBT-1. The experimental data revealed that SBT-1 possessed the ability to ferment relatively high glucose or sucrose concentrations to D-lactic acid without obvious catabolite repression and byproduct formation compared to the two reference strains. In draft genome sequencing of S. terrae SBT-1, the results provided here can promote further study to overcome the current limitations for the industrial-scale production of D-lactic acid.


Assuntos
Bacillales , Fermentação , Genoma Bacteriano , Ácido Láctico , Açúcares , Bacillales/genética , Genoma Bacteriano/genética , Glucose/metabolismo , Ácido Láctico/metabolismo , Açúcares/metabolismo
8.
Antonie Van Leeuwenhoek ; 114(7): 1043-1057, 2021 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-33913068

RESUMO

A cellulase producing novel bacterial strain VR-M41T was isolated from an open-air vegetable and fruit market. Cells are found to be rod-shaped, endospore forming, positive for Gram's stain and negative for catalase, oxidase and urease. Strain VR-M41T was halotolerant (upto 8.0% NaCl, w/v), motile and facultative anaerobe. It grew at wide range of pH (6.0-10.0) and temperatures (20-40 °C). Strain VR-M41T produced three isozymes of Carboxymethylcellulase. The 16S rRNA gene sequence of strain VR-M41T was 97.3% similar to both Saccharibacillus kuerlensis DSM 22868T and Saccharibacillus sacchari DSM 19268T, and less than 96.4% with the rest of the valid species of the genus Saccharibacillus. Whole-genome ANI, dDDH and genome phylogenetic tree analysis revealed that strain VR-M41T significantly differed from Saccharibacillus kuerlensis DSM 22868T and Saccharibacillus sacchari DSM 19268T (ANI 79.6-79.7% and dDDH 23.1%). The strain comprised of MK-7 and anteiso-C 15:0 (42.2%) as predominant isoprenoid quinone and fatty acid respectively. Major polar lipids were found to be diphosphatidylglycerol, phosphatidylglycerol and phosphatidylethanolamine. The draft genome of strain VR-M41T consisted of 5,386,426 base pairs with 5103 annotated genes, out of which 2147 corresponded to hypothetical proteins and 2956 with functional assignments. Pan-genome analysis revealed the presence of 2998 core genes, 828 accessory genes, and 1131 unique genes of Saccharibacillus. Strain VR-M41T produced antimicrobials against Staphylococcus aureus, Streptococcus pneumoniae, Micrococcus luteus and Shigella flexneri. Significant phenotypic and genotypic differentiating characteristics from closely related species, indicated that strain VR-M41T is a novel species of the genus Saccharibacillus, for which the name Saccharibacillus alkalitolerans sp. nov., is proposed. The type strain is VR-M41T (= KCTC 43183T=NBRC 114337T).


Assuntos
Anti-Infecciosos , Celulases , Bacillales , Técnicas de Tipagem Bacteriana , Composição de Bases , DNA Bacteriano/genética , Ácidos Graxos/análise , Genômica , Isoenzimas , Hibridização de Ácido Nucleico , Fosfolipídeos , Filogenia , RNA Ribossômico 16S/genética , Análise de Sequência de DNA
9.
Arch Microbiol ; 203(5): 2445-2451, 2021 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-33666688

RESUMO

A Gram-positive, aerobic, endospore-forming, rod-shaped bacterial strain, CAU 1483 T, was isolated from tidal-flat mud in the Republic of Korea. It grew optimally at 30 °C, in a pH 7.0 medium with 2% (w/v) NaCl. Phylogenetic analysis based on the 16S rRNA gene sequence indicated that strain CAU 1483 T formed a separate clade within Paenibacillaceae together with members of the genus Cohnella. Strain CAU 1483 T exhibited the highest 16S rRNA gene sequence similarity (97.1%) to C. candidum 18JY8-7 T. Whole genome of strain CAU 1483 T was 4.29 Mb in size with a 53.7 mol% G + C content, and included 4046 coding sequences and included 4046 coding sequences, some of which associated with stress response. The average nucleotide identity and digital DNA-DNA hybridization similarity between strain CAU 1483 T and related members of the genus Cohnella were 71.8-74.9% and 22.6-33.9%, respectively. The major respiratory quinone present in this strain was menaquinone-7. Strain CAU 1483 T contained anteiso-C15:0 and iso-C16:0 as the major fatty acids, while its polar lipids consisted of phosphatidylglycerol, phosphatidylethanolamine, diphosphatidylglycerol, lysyl-phosphatidylglycerol, phosphatidylcholine, three unidentified aminophospholipids, two unidentified lipids and an unidentified phospholipid. Peptidoglycan type was A1γ meso-Dpm. On the basis of taxonomic characterization, strain CAU 1483 T constitutes a novel species, for which the name Cohnella pontilimi sp. nov. is proposed. The type strain of this novel species is CAU 1483 T (= KCTC 43047 T = NBRC 113953 T).


Assuntos
Bacillales/classificação , Sedimentos Geológicos/microbiologia , Bacillales/química , Bacillales/fisiologia , DNA Bacteriano/genética , Ácidos Graxos/química , Genoma Bacteriano/genética , Peptidoglicano/química , Fosfolipídeos/química , Filogenia , RNA Ribossômico 16S/genética , República da Coreia , Análise de Sequência de DNA , Vitamina K 2/análogos & derivados
10.
Animal ; 15(2): 100112, 2021 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-33573975

RESUMO

Silkworm (Bombyx mori L.) pupae are a by-product derived from silk production, which is often treated as waste and thus discarded: this can cause serious environmental problems and a loss of nutrients. Silkworm pupae are a rich source of protein and lipids, and the resulting protein meal can provide promising outcomes as livestock feed, notably for monogastric species. However, one possible issue that needs to be considered is the possible implication of the 1-Deoxynojirimycin (1-DNJ), a bio-compound of the silkworm that impairs glucose absorption, in poultry nutrition. Therefore, the present study evaluated the effect of the dietary inclusion of full-fat or defatted silkworm pupa meal (SWM) on the apparent digestibility of nutrients, feed choice and faecal microbiome in meat-producing quails. For the digestibility trial, a total of thirty-three 27-day-old Japanese quails (Coturnix coturnix japonica) were individually housed in digestibility cages and received three experimental diets: a control diet (control, commercial feed for fattening quails), and two other diets containing the 12.5% of either a full-fat SWM (SWM-FULL) or a defatted SWM (SWM-DEF). Subsequently, twenty-seven 33-day-old quails were simultaneously provided with Control, SWM-FULL and SWM-DEF diets for a 10-day feed choice trial. The results of the digestibility trial showed that the DM intake and excreta production were higher in both SWM groups than in the Control one (P < 0.001). The apparent digestibility of DM, organic matter, CP, ether extract, starch and energy was lower in both SWM groups than in the control group (P < 0.001), suggesting the possible implication of chitin and 1-DNJ. The feed choice test showed that quails preferred the Control diet (P < 0.001). From the microbiome analysis of the excreta, families such as Streptococcaceae (P < 0.05), Rikenellaceae and Eubacteriaceae (P < 0.01) and taxa at species level such as Lactobacillus delbrueckii (P < 0.05), Aneurinibacillus thermoaerophilus and Bacillus thermoamylovorans (P < 0.01) scored higher in SWM-FULL quails than in SWM-DEF and Control treatments. The present study demonstrated that a successful dietary inclusion of SWM for fattening quails needs to overcome the digestive criticalities caused by the of presence specific bio-compounds, namely chitin and 1-DNJ.


Assuntos
Ração Animal , Bombyx , Microbiota , Ração Animal/análise , Fenômenos Fisiológicos da Nutrição Animal , Animais , Bacillales , Bacillus , Coturnix , Dieta/veterinária , Digestão , Nutrientes , Pupa , Codorniz
11.
Antonie Van Leeuwenhoek ; 114(1): 37-44, 2021 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-33392934

RESUMO

A Gram-stain positive, aerobic, motile, rod-shaped bacterium designated as strain CBP-2801T was isolated as a contaminant from a culture containing maize callus in Peoria, Illinois, United States. The strain is unique relative to other Cohnella species due to its slow growth and reduced number of sole carbon sources. Phylogenetic analysis using 16S rRNA indicated that strain CBP-2801T is a Cohnella bacterium and showed the highest similarity to Cohnella xylanilytica (96.8%). Genome-based phylogeny and genomic comparisons based on average nucleotide identity confirmed the strain to be a novel species of Cohnella. Growth occurs at 15-45 °C (optimum 40 °C), pH 5-7 (optimum pH 6) and with 0-1% NaCl. The predominant fatty acids are anteiso-15:0 and 18:1 ω6c. Genome mining for secondary metabolites identified a putative biosynthetic cluster that encodes for a novel lasso peptide. In addition, this study contributes five new genome assemblies of type strains of Cohnella species, a genus with less than 30% of the type strains sequenced. The DNA G + C content is 58.7 mol %. Based on the phenotypic, phylogenetic and biochemical data strain CBP-2801T represents a novel species, for which the name Cohnella zeiphila sp. nov. is proposed. The type strain is CBP-2801T (= DSM 111598 = ATCC TSD-230).


Assuntos
Fosfolipídeos , Zea mays , Bacillales , Técnicas de Tipagem Bacteriana , Composição de Bases , DNA Bacteriano/genética , Ácidos Graxos/análise , Fosfolipídeos/análise , Filogenia , RNA Ribossômico 16S/genética , Análise de Sequência de DNA
12.
Environ Pollut ; 272: 116009, 2021 Mar 01.
Artigo em Inglês | MEDLINE | ID: mdl-33257150

RESUMO

Biochar has been proven as a soil amendment to improve soil environment. However, mechanistic understanding of biochar on soil physical properties and microbial community remains unclear. In this study, a wood biochar (WB), was incorporated into a highly weathered tropical soil, and after 1 year the in situ changes in soil properties and microbial community were evaluated. A field trial was conducted for application of compost, wood biochar, and polyacrylamide. Microstructure and morphological features of the soils were characterized through 3D X-ray microscopy and polarized microscopy. Soil microbial communities were identified through next-generation sequencing (NGS). After incubation, the number of pores and connection throats between the pores of biochar treated soil increased by 3.8 and 7.2 times, respectively, compared to the control. According to NGS results, most sequences belonged to Anaerolinea thermolimosa, Caldithrix palaeochoryensis, Chthoniobacter flavus, and Cohnella soli. Canonical correlation analysis (CCA) further demonstrated that the microbial community structure was determined by inorganic N (IN), available P (AP), pH, soil organic C (SOC), porosity, bulk density (BD), and aggregate stability. The treatments with co-application of biochar and compost facilitated the dominance of Cal. palaeochoryensis, Cht. flavus, and Coh. soli, all of which promoted organic matter decomposition and ammonia oxidation in the soil. The apparent increases in IN, AP, porosity, and SOC caused by the addition of biochar and compost may be the proponents of changes in soil microbial communities. The co-application of compost and biochar may be a suitable strategy for real world biochar incorporation in highly weathered soil.


Assuntos
Carvão Vegetal , Solo , Bacillales , Bactérias , Chloroflexi , Microbiologia do Solo , Verrucomicrobia
13.
Pest Manag Sci ; 77(4): 2019-2028, 2021 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-33342024

RESUMO

BACKGROUND: Bacillus thuringiensis (Bt) is a Gram-positive bacterium that synthesizes specific protein toxins, which can be exploited for control of various insect pests, including Diatraea saccharalis, a lepidopteran that severely damages sugarcane crops. Although studies have described the effects of Bt in the larval phases of D. saccharalis, few have examined its effect on insect eggs. Herein, we studied the entomopathogenic potential of Bacillus thuringiensis serovar Aizawai GC-91 (Bta) during D. saccharalis embryo development with the aim of understanding the entomopathogenic mechanism and developing new biological control techniques for target insects. RESULTS: Bta concentrations of 5, 10 and 20 g L-1 demonstrated the strongest bioactivity, reducing D. saccharalis egg viability by 28.69%, 33.91% and 34.98%, respectively. The lethal concentrations (LCs) were estimated as: LC50 = 28.07 g L-1 (CI 95% = 1.89-2.38) and LC90 = 65.36 g L-1 (CI 95% = 4.19-5.26). Alterations in egg coloration, melanization and granule accumulation were observed at 24 h, persisting until 144 h. The embryo digestive systems were severely damaged, including narrowing of the intestinal lumen, vesiculations and degenerated cells, causing embryonic death. CONCLUSION: The toxicity caused by Bta in D. saccharalis embryos demonstrated its potential as a biological control agent and as a sustainable alternative for integrated management of D. saccharalis infestation. © 2020 Society of Chemical Industry.


Assuntos
Bacillaceae , Bacillales , Bacillus thuringiensis , Lepidópteros , Mariposas , Animais , Toxinas de Bacillus thuringiensis , Proteínas de Bactérias , Endotoxinas , Proteínas Hemolisinas , Larva , Controle Biológico de Vetores
14.
Microbiol Res ; 242: 126614, 2021 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-33045681

RESUMO

The presence of biosurfactants produced by a Bacillus strain in corn steep liquor (CSL), a wastewater stream of the corn milling process, has been recently discovered. However, the species responsible for their production has not been identified at the moment. Therefore, in this work, the Bacillus strain isolated from CSL, with capacity to produce biosurfactants, was subjected to amplification and sequence analysis of the 16S rRNA, being identified as Aneurinibacillus aneurinilyticus. This strain has been proved to be endospore forming and thermophile, what would explain its presence in the commercial CSL. It was observed that the strain under evaluation has the ability to produce both cell-bound and extracellular biosurfactant extracts, which were characterized in this work. The electrospray ionization mass spectrometry (ESI) analysis of the biosurfactant extracts revealed that the extracellular biosurfactant produced by Aneurinibacillus aneurinilyticus is composed by a mixture of lipopeptides, containing C16 and C18 fatty acids and amino acids, including valine, phenylalanine, proline, cysteine, histidine, aspartic acid/asparagine, alanine, glycine, leucine/isoleucine, with biomarkers between 1025-458 m/z. Conversely, the cell-bound biosurfactant extract produced by Aneurinibacillus aneurinilyticus was composed by the cyclic decapeptide gramicidin S, with a characteristic peak at 571 m/z, and lipopeptides with characteristic peaks between 1034-705 m/z, containing alanine, glycine, cysteine, serine, proline, aspartic acid/asparagine, similarly to the amino acid sequence of the extracellular biosurfactant extract.


Assuntos
Bacillales/isolamento & purificação , Bacillales/metabolismo , Tensoativos/química , Tensoativos/metabolismo , Zea mays/microbiologia , Aminoácidos/análise , Bacillales/genética , Bacillus/classificação , Bacillus/genética , Bacillus/isolamento & purificação , Ácidos Graxos/análise , Gramicidina/metabolismo , Lipopeptídeos/análise , RNA Ribossômico 16S/genética , Espectrometria de Massas por Ionização por Electrospray
15.
Electron. j. biotechnol ; 47: 83-88, sept. 2020. graf, ilus
Artigo em Inglês | LILACS | ID: biblio-1253097

RESUMO

BACKGROUND: L-tert-Leucine has been widely used in pharmaceutical, chemical, and other industries as a vital chiral intermediate. Compared with chemical methods, enzymatic methods to produce L-tert-leucine have unparalleled advantages. Previously, we found a novel leucine dehydrogenase from the halophilic thermophile Laceyella sacchari (LsLeuDH) that showed good thermostability and great potential for the synthesis of L-tertleucine in the preliminary study. Hence, we manage to use the LsLeuDH coupling with a formate dehydrogenase from Candida boidinii (CbFDH) in the biosynthesis of L-tert-leucine through reductive amination in the present study. RESULT: The double-plasmid recombinant strain exhibited higher conversion than the single-plasmid recombinant strain when resting cells cultivated in shake flask for 22 h were used. Under the optimized conditions, the double-plasmid recombinant E. coli BL21 (pETDute-FDH-LDH, pACYCDute-FDH) transformed 1 mol·L-1 trimethylpyruvate (TMP) completely into L-tert-leucine with greater than 99.9% ee within 8 h. CONCLUSIONS: The LsLeuDH showed great ability to biosynthesize L-tert-leucine. In addition, it provided a new option for the biosynthesis of L-tert-leucine.


Assuntos
Leucina Desidrogenase/metabolismo , Bacillales/enzimologia , Leucina/biossíntese , Temperatura , Proteínas Recombinantes , Escherichia coli , Concentração de Íons de Hidrogênio
16.
Carbohydr Polym ; 247: 116714, 2020 Nov 01.
Artigo em Inglês | MEDLINE | ID: mdl-32829841

RESUMO

Enzymatic transformation of xylans into renewable fuels and value-added products is mediated by xylanases. Here we describe the biochemical and X-ray structural characterization of Thermobacillus composti GH10 xylanase (TcXyn10A) at 2.1 Å resolution aiming to unravel details of its recognition of glucurono- and arabinoxylan at a molecular level. TcXyn10A improves the efficiency of pretreated lignocellulosic biomass hydrolysis by a commercial enzyme cocktail causing a 15.35 % increase in xylose release and 4.38 % glucose release after 24 h of reaction. The enzyme releases predominantly xylobiose and xylotriose, as well as MeGlcA3 × 3 (from beechwood glucuronoxylan) and a range of decorated xylooligosaccharides (XOS) from rye arabinoxylan, with Ara2 × 2 being the major product. The enzyme liberates XOS with the yields of 29.09 % for beechwood glucuronoxylan and 16.98 % for rye arabinoxylan. Finally, TcXyn10A has a high thermal stability, halotolerance, and resistance to ethanol, biochemical properties that can be desirable for a number of industrial applications.


Assuntos
Bacillales/enzimologia , Endo-1,4-beta-Xilanases/metabolismo , Xilanos/química , Hidrólise , Especificidade por Substrato , Xilanos/metabolismo
17.
Int J Biol Macromol ; 163: 1369-1374, 2020 Nov 15.
Artigo em Inglês | MEDLINE | ID: mdl-32758598

RESUMO

Xylan and pectin are major structural components of plant cell walls. There are two independent catabolic pathways for xylan and pectin. UxaE bridges these two pathways by reversibly epimerizing D-fructuronate and D-tagaturonate. The crystal structure of UxaE from Cohnella laeviribosi (ClUxaE) shows a core scaffold of TIM-barrel with a position-changing divalent metal cofactor. ClUxaE has the flexible metal-coordination loop to allow the metal shift and the extra domains to bind a phosphate ion in the active site, which are important for catalysis and substrate specificity. Elucidation of the structure and mechanism of ClUxaE will assist in understanding the catalytic mechanism of UxaE family members, which are useful for processing both xylan and pectin-derived carbohydrates for practical and industrial purposes, including the transformation of agricultural wastes into numerous valuable products.


Assuntos
Bacillales/química , Catálise/efeitos dos fármacos , Racemases e Epimerases/química , Açúcares/química , Carboidratos/química , Domínio Catalítico/efeitos dos fármacos , Metais/química , Pectinas/química , Fosfatos/química , Especificidade por Substrato , Xilanos/química
18.
Artigo em Inglês | MEDLINE | ID: mdl-32664813

RESUMO

Alkaline proteases having activity and stability at alkaline pH possess a large variety of applications in many industries. Growing renewed interest urges the need to find a single alkaline protease with promising properties to be used in different industrial processes. Herein, alkaline proteases produced through fermentation of cheap and easily available organic municipal solid wastes by Bacillus subtilis AKAL7 and Exiguobacterium indicum AKAL11 were purified to investigate their kinetic and thermodynamic parameters, detergent compatibility, dehairing and feather-degrading capability. Sodium dodecyl sulfate polyacrylamide gel electrophoresis revealed that the purified protease from B. subtilis and E. indicum had molecular mass of ∼45 and 75 kDa, respectively. The protease from B. subtilis and E. indicum showed highest activity at 55 and 50 °C having low K m 1.17 and 0.567 mg/mL and high V max 416.67 and 333.33 µmole/min, respectively. The activation energy and temperature quotient of protease from B. subtilis and E. indicum were 26.52 and 65.75 kJ/mole, and 1.0004 and 1.0003 at 20-55 and 20-50 °C, respectively. Thermodynamics analysis revealed the formation of more ordered enzyme-substrate complexes along with spontenity of enzyme reaction. The protease from E. indicum exhibited better compatibility at higher concentration of detergents compared to that from B. subtilis. However, both proteases could retain more than 80% of the activity in the presence of 0.1% commercial laundry detergents. The purified protease from the both sources could degrade almost 90% of barbs and 40% of dry weight of the native feather and that from E. indicum could dehair cow skin. Results reported herein suggest that the alkaline protease from B. subtilis AKAL7 and E. indicum AKAL11 has biotechnological implications in detergent, leather and poultry feather processing industries.


Assuntos
Bacillales/enzimologia , Bacillus subtilis/enzimologia , Proteínas de Bactérias/metabolismo , Endopeptidases/metabolismo , Resíduos Sólidos , Animais , Detergentes/química , Estabilidade Enzimática , Exiguobacterium , Plumas , Fermentação , Cinética , Peso Molecular , Temperatura
19.
PLoS One ; 15(6): e0234958, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32574185

RESUMO

Proteases play an essential role in living organisms and represent one of the largest groups of industrial enzymes. The aim of this work was recombinant production and characterization of a newly identified thermostable protease 1147 from thermophilum indigenous Cohnella sp. A01. Phylogenetic tree analysis showed that protease 1147 is closely related to the cysteine proteases from DJ-1/ThiJ/PfpI superfamily, with the conserved catalytic tetrad. Structural prediction using MODELLER 9v7 indicated that protease 1147 has an overall α/ß sandwich tertiary structure. The gene of protease 1147 was cloned and expressed in Escherichia coli (E. coli) BL21. The recombinant protease 1147 appeared as a homogenous band of 18 kDa in SDS-PAGE, which was verified by western blot and zymography. The recombinant protein was purified with a yield of approximately 88% in a single step using Ni-NTA affinity chromatography. Furthermore, a rapid one-step thermal shock procedure was successfully implemented to purify the protein with a yield of 73%. Using casein as the substrate, Km, and kcat, kcat/Km values of 13.72 mM, 3.143 × 10-3 (s-1), and 0.381 (M-1 S-1) were obtained, respectively. The maximum protease activity was detected at pH = 7 and 60°C with the inactivation rate constant (kin) of 2.10 × 10-3 (m-1), and half-life (t1/2) of 330.07 min. Protease 1147 exhibited excellent stability to organic solvent, metal ions, and 1% SDS. The protease activity was significantly enhanced by Tween 20 and Tween 80 and suppressed by cysteine protease specific inhibitors. Docking results and molecular dynamics (MD) simulation revealed that Tween 20 interacted with protease 1147 via hydrogen bonds and made the structure more stable. CD and fluorescence spectra indicated structural changes taking place at 100°C, very basic and acidic pH, and in the presence of Tween 20. These properties make this newly characterized protease a potential candidate for various biotechnological applications.


Assuntos
Bacillales/enzimologia , Proteínas de Bactérias/química , Peptídeo Hidrolases/química , Proteínas de Bactérias/isolamento & purificação , Proteínas de Bactérias/ultraestrutura , Clonagem Molecular , Ensaios Enzimáticos , Estabilidade Enzimática , Concentração de Íons de Hidrogênio , Simulação de Dinâmica Molecular , Peso Molecular , Peptídeo Hidrolases/isolamento & purificação , Peptídeo Hidrolases/ultraestrutura , Proteínas Recombinantes/química , Proteínas Recombinantes/isolamento & purificação , Proteínas Recombinantes/ultraestrutura , Especificidade por Substrato
20.
Ecotoxicol Environ Saf ; 201: 110860, 2020 Sep 15.
Artigo em Inglês | MEDLINE | ID: mdl-32563162

RESUMO

The biodegradation of naphthalene using low-density polyethylene (LDPE) immobilized Exiguobacterium sp. RKS3 (MG696729) in a packed bed bioreactor (PBBR) was studied. The performance of a continuous PBBR was evaluated at different inlet flow rates (IFRs) (20-100 mL/h) under 64 days of operation. The maximum naphthalene removal efficiency (RE) was found at low IFR, and it further decreased with increasing IFRs. In a continuous PBBR, the external mass transfer (EMT) aspect was analysed at various IFRs, and experimental data were interrelated between Colburn factor (JD) and Reynolds number (NRe) as [Formula: see text] . A new correlation [Formula: see text] was obtained to predict the EMT aspect of naphthalene biodegradation. Andrew-Haldane model was used to evaluate the bio-kinetic parameters of naphthalene degradation, and kinetic constant νmax, Js, and Ji were found as 0.386 per day, 13.6 mg/L, and 20.54 mg/L, respectively.


Assuntos
Reatores Biológicos/microbiologia , Naftalenos/análise , Poluentes Químicos da Água/análise , Purificação da Água/métodos , Bacillales/crescimento & desenvolvimento , Bacillales/metabolismo , Biodegradação Ambiental , Células Imobilizadas/microbiologia , Cinética , Polietileno/química
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