Metagenomic data mining of the Nellore cattle rumen microbiota identified a new bifunctional enzyme, endo-1,4-β-xylanase/esterase, which was subsequently overexpressed in E. coli BL21 (DE3). This ...enzyme was stable at pH intervals of 5 to 6.5 and temperatures between 30 and 45 °C, and under the test conditions, it had a V
of 30.959 ± 2.334 µmol/min/mg, K
of 3.6 ± 0.6 mM and k
of 2.323 ± 175 s
. Additionally, the results showed that the enzyme is tolerant to NaCl and organic solvents and therefore is suitable for industrial environments. Xylanases are widely applicable, and the synergistic activity of endo-1,4-β-xylanase/esterase in a single molecule will improve the degradation efficiency of heteroxylans via the creation of xylanase binding sites. Therefore, this new molecule has the potential for use in lignocellulosic biomass processing and as an animal feed food additive and could improve xylooligosaccharide production efficiency.
The aminopeptidase gene from Mesorhizobium SEMIA3007 was cloned and overexpressed in Escherichia coli. The enzyme called MesoAmp exhibited optimum activity at pH 8.5 and 45 °C and was strongly ...activated by Co
and Mn
. Under these reaction conditions, the enzyme displayed K
and k
values of 0.2364 ± 0.018 mM and 712.1 ± 88.12 s
, respectively. Additionally, the enzyme showed remarkable stability in organic solvents and was active at high concentrations of NaCl, suggesting that the enzyme might be suitable for use in biotechnology. MesoAmp is responsible for 40% of the organism's aminopeptidase activity. However, the enzyme's absence does not affect bacterial growth in synthetic broth, although it interfered with biofilm synthesis and osmoregulation. To the best of our knowledge, this report describes the first detailed characterization of aminopeptidase from Mesorhizobium and suggests its importance in biofilm formation and osmotic stress tolerance. In summary, this work lays the foundation for potential biotechnological applications and/or the development of environmentally friendly technologies and describes the first solvent- and halo-tolerant aminopeptidases identified from the Mesorhizobium genus and its importance in bacterial metabolism.
Esterases catalyze the cleavage and formation of ester bonds and are members of the diverse family of α/β hydrolase fold. They are useful in industries from different sectors, such as food, ...detergent, fine chemicals, and biofuel production. In a previous work, 30 positive clones for lipolytic activity were identified from a metagenomic library of a microbial consortium specialized in diesel oil degradation. In this study, a putative gene encoding an esterase/lipase, denominated
est8
, has been cloned and the corresponding protein expressed recombinantly, purified to homogeneity and characterized functional and structurally. We show that the protein codified by
est8
gene, denominated Est8, is an alkaline esterase with high catalytic efficiency against
p
-nitrophenyl acetate and stable in the presence of up to 10% dimethyl sulfoxide. The three-dimensional structure of Est8 was determined at 1.85-Ǻ resolution, allowing the characterization of the substrate-binding pocket and features that rationalize the preference of Est8 for short-chain substrates. In an attempt to increase the size of ligand-binding pocket and enzyme activity against distinct substrates of long chain, we mutated two residues (Met
213
and Phe
217
) that block the substrate channel. A small increase in the reaction velocity for
p
-nitrophenyl butyrate and
p
-nitrophenyl valerate hydrolysis was observed. Activity against
p
-nitrophenyl acetate was reduced. The functional and structural characterization of Est8 is explored in comparison with orthologues.
After being isolated from a sugarcane pile, the bacterium Chitinophaga sp. CB10 demonstrated to be a rich source of carbohydrases, with 350 predicted CAZyme domains. CB10 was able to grow on ...carbohydrates of different structural complexities: glucose, carboxymethylcellulose, corn starch, galactomannan, Aloe vera gum and sugarcane bagasse. The sugarcane bagasse is a rich source of complex polymers, and the diversity of metabolites released by its enzymatic hydrolysis has an important role for green chemistry, including minority pathways such as the degradation of mannan conjugates. In this sense, CB10 demonstrated considerable levels of gene expression for mannanases, and was stable for a period of 96-144 hours in the presence of sugarcane bagasse as sole carbon source. The bacterium showed respectively 4.8x and 5.6x expression levels for two genes predicted for GH2 β-mannosidase: one located within a gene cluster identified as "polysaccharide utilization loci" (PUL), and another a classic β-mannosidase. These enzymes shared less than 45% of identity with enzymes characterized from the genus Chitinophaga belonging to the phylum Bacteroidetes. The degree of novelty-as demonstrated by the low identity with previously characterized enzymes; the remarkable capability to grow in different substrates; mannanase activity, evidenced by the release of residual oligosaccharides in the cultivation with galactomannan (HPLC-RID, 12.3 mMol); associated to the ability of mannanases expression in a low concentration of inductor conditions (sugarcane bagasse, 0.2%) indicate the high potential for the application of CB10 as a source of enzymes in the production of oligosaccharides from biomass. This capacity might prove to be very valuable for the biorefinery process of pre-biotic precursors and other functional oligosaccharides focused on the food and pharmaceutical industries.
Members of subdivision 1 of the phylum Acidobacteria were grown at different pH values in a new medium formulation named PSYL 5, which includes sucrose as a carbon source and other compounds (such as ...KH2PO4 and MgSO4.7H2O). Growth rate was nearly constant at pH 5.0 and declined at pH 3–4 and 6–7. However, it was found that effects involving good carbon/nitrogen ratios and pH on the growth of the members of Acidobacteria subdivision 1 were significant, and the strongest effect of these conditions was at pH 5.0. In addition, incubation time of 48, 72, 96 and 120 h was shorter than that described previously for members of Acidobacteria subdivision 1 on solid laboratory media.
New medium formulation named PSYL 5 includes sucrose, as a carbon source, and other compounds (such as KH2PO4 and MgSO4.7H2O) for members of Acidobacteria subdivision 1.
The influence of bivalent copper (Cu
2+
) and hexavalent chromium (Cr
6+
) ions on exopolysaccharide (EPS) and polyhydroxybutyrate (PHB) production by the bacterium
Rhizobium tropici
LBMP-C01 was ...investigated. The partial structure of both biopolymers produced by rhizobia were characterized, and the quantification of
exo
R and
exo
Z gene expression by real-time polymerase chain reaction (Real-Time PCR) was also investigated. Results showed that the supplementation with Cu
2+
for 144 h induced an increase in the production of EPS by 48% and PHB by 46.66% compared to the control. Expression studies revealed that the expressed
exo
R and e
xo
Z genes were affected by metal ion treatment. It can be stated that this particular rhizobial strain is a future potential candidates to produce super adsorbent quite useful for soil bioremediation.
Evidence based on genomic sequences is extremely important to confirm the phylogenetic relationships within the Rhizobium group. SEMIA3007 was analyzed within the Mesorhizobium groups to define the ...underlying causes of taxonomic identification. We previously used biochemical tests and phenotypic taxonomic methods to identify bacteria, which can lead to erroneous classification. An improved understanding of bacterial strains such as the Mesorhizobium genus would increase our knowledge of classification and evolution of these species.
In this study, we sequenced the complete genome of SEMIA3007 and compared it with five other Mesorhizobium and two Rhizobium genomes. The genomes of isolated SEMIA3007 showed several orthologs with M. huakuii, M. erdmanii and M. loti. We identified SEMIA3007 as a Mesorhizobium by comparing the 16S rRNA gene and the complete genome.
Our ortholog, 16S rRNA gene and average nucleotide identity values (ANI) analysis all demonstrate SEMIA3007 is not Rhizobium leguminosarum bv. viceae. The results of the phylogenetic analysis clearly show SEMIA3007 is part of the Mesorhizobium group and suggest a reclassification is warranted.
Lipolytic enzymes have attracted attention from a global market because they show enormous biotechnological potential for applications such as detergent production, leather processing, cosmetics ...production, and use in perfumes and biodiesel. Due to the intense demand for biocatalysts, a metagenomic approach provides methods of identifying new enzymes. In this study, an esterase designated as Est16 was selected from 4224 clones of a fosmid metagenomic library, revealing an 87% amino acid identity with an esterase/lipase (accession number ADM63076.1) from an uncultured bacterium. Phylogenetic studies showed that the enzyme belongs to family V of bacterial lipolytic enzymes and has sequence and structural similarities with an aryl-esterase from Pseudomonas fluorescens and a patented Anti-Kazlauskas lipase (patent number US20050153404). The protein was expressed and purified as a highly soluble, thermally stable enzyme that showed a preference for basic pH. Est16 exhibited activity toward a wide range of substrates and the highest catalytic efficiency against p-nitrophenyl butyrate and p-nitrophenyl valerate. Est16 also showed tolerance to the presence of organic solvents, detergents and metals. Based on molecular modeling, we showed that the large alpha-beta domain is conserved in the patented enzymes but not the substrate pocket. Here, it was demonstrated that a metagenomic approach is suitable for discovering the lipolytic enzyme diversity and that Est16 has the biotechnological potential for use in industrial processes.
Hypertension and diabetes mellitus are the second and third highest leading causes of disability-adjusted life-years (DALY), respectively, in Brazil. The clinical outcomes of chronic diseases are ...influenced by various factors. Therefore, there is a need for multifaceted interventions to achieve a decrease in the rate of DALY, with a better control of these diseases.
To verify whether sustainable long-term interventions, such as health worker training and provision of health education to the patients, contribute to health improvements in patients with hypertension and diabetes from rural communities.
Over a 6 month period, educational and medical interventions were provided to optimize the treatment of hypertension and diabetes. Furthermore, blood pressure and glycated hemoglobin (HbA1c) measurements were taken at baseline and after the interventions.
The monitored hypertensive patients (
= 276) had a reduction of 13.4 mmHg (
= 0.021) and 5.8 mmHg (
< 0.001) in mean systolic and diastolic blood pressure, respectively. Diabetic patients who were followed-up (
= 71) achieved a 0.55% (
= 0.185) reduction in HbA1c level. The desired blood pressure level (<140/90 mmHg) was achieved in 38.8% of patients with hypertension, whereas the desired level of HbA1c (<7.0% for adults and <8.0% for the elderly) was achieved in 16.9% of patients with diabetes; in addition, 38.0% had a reduction of HbA1c of at least 1%.
The results showed that the interventions improved the blood pressure and HbA1c levels in patients with hypertension and diabetes from rural communities in a municipality in Northeast Brazil.
The aim of the present study was to analyse the genetic and pathogenic variability of Colletotrichum spp. isolates from various organs and cultivars of mango with anthracnose symptoms, collected from ...different municipalities of São Paulo State, Brazil. Colletotrichum gloeosporioides isolates from symptomless citrus leaves and C. acutatum isolates from citrus flowers with post‐bloom fruit drop symptoms were included as controls. Sequencing of the ITS region allowed the identification of 183 C. gloeosporioides isolates from mango; only one isolate was identified as C. acutatum. amova analysis of ITS sequences showed larger genetic variability among isolates from the same municipality than among those from different populations. fAFLP markers indicated high levels of genetic variability among the C. gloeosporioides isolates from mango and no correlation between genetic variability and isolate source. Only one C. gloeosporioides mango isolate had the same genotype as the C. gloeosporioides isolates from citrus leaves, as determined by ITS sequencing and fAFLP analysis. Pathogenicity tests revealed that C. gloeosporioides and C. acutatum isolates from either mango or citrus can cause anthracnose symptoms on leaves of mango cvs Palmer and Tommy Atkins and blossom blight symptoms in citrus flowers. These outcomes indicate a lack of host specificity of the Colletotrichum species and suggest the possibility of host migration.
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