Despite being one of the most isolated regions in the world, Antarctica is at risk of increased contamination with potentially toxic elements and other toxic chemicals through anthropogenic ...interventions. In this study, a psychrotolerant bacterium was isolated using the lake water collected from Ardley Island (Antarctica), which can grow at temperatures between 4 and 30 °C and pH values between 6.0 and 9.0. The isolate, named AC, had protease, amylase, and lipase activities with no NaCl tolerance and could degrade 1–5% diesel fuel. Multilocus sequence analysis (MLSA) using 16S rRNA, gyrB, tuf, and rpoD genes resulted in 92.91–98.6% sequence similarities between the isolate AC and other Flavobacterium spp. Whole genome analysis indicated that the genome length of Flavobacterium sp. AC is 5.8 Mbp with a GC content of 34.04% and 1274 genes predicted. The strain AC branched independently from other Flavobacterium spp. in the phylogenetic and phylogenomic trees and ranked a new species named Flavobacterium aziz-sancarii. Genome mining identified several cold-inducible genes, including stress-associated genes such as cold-shock proteins, chaperones, carotenoid biosynthetic genes, or oxidative-stress response genes. In addition, virulence, gliding motility, and biofilm-related genes were determined. Its genome contains 35 and 88 open-reading frames related to potentially toxic element and antibiotic resistance, respectively. F. aziz-sancarii showed a remarkable tolerance of Cr and Ni, with minimal inhibitory concentration values of 2.88 and 2.81 mM, respectively. Pb, Cu, and Zn exposure resulted in moderate toxicity (2.14–2.41 mM), while Cd showed the highest inhibitory effect in bacterial growth (0.74 mM). Antibiotic susceptibility testing indicated multidrug-resistant phenotype in correlation to in silico prediction of antibiotic resistance genes. Overall, our results contribute to biodiversity of Antarctica and provide new insights into resistome profile of Antarctic microorganisms. Additionally, the diesel degradation feature of F. aziz-sancarii offers potential use for the bioremediation of hydrocarbon-contaminated polar ecosystems.
Display omitted
•Flavobacterium aziz-sancarii can degrade diesel and produce hydrolytic enzymes•It displayed multidrug resistant phenotype and potentially toxic element tolerance•Various genes related to resistome, virulence, motility, and biofilm were detected•A good correlation between genome content and phenotype was revealed
Display omitted
•Transporters and transcriptional regulators were the main DEGs in the mutant strain.•Siderophore-related genes were upregulated in the absence of iron in the mutant.•Siderophore ...production was enhanced in the mutant in iron scarcity.•Antibiotic production by the mutant was improved in iron-containing media.
Iron homeostasis is strictly regulated by complex cascades connected with secondary metabolism in bacteria. Ferric uptake regulators ('Fur's), siderophores, efflux systems, and two-component signal transduction systems are the leading players in response stimuli. However, these regulatory mechanisms remain to be elucidated in Streptomyces clavuligerus. Our study focused on unraveling a possible role of SCLAV_3199 which encodes a Fur family transcriptional regulator, particularly in iron regulation and at the global level in this species. We deleted the SCLAV_3199 gene in S. clavuligerus and compared gene expression differences with the wild-type strain based on iron availability by RNA-seq. We found a potential regulatory effect of SCLAV_3199 on many transcriptional regulators and transporters. Besides, the genes encoding iron sulfur binding proteins were overexpressed in the mutant in the presence of iron. Notably, catechol (SCLAV_5397), and hydroxamate-type (SCLAV_1952, SCLAV_4680) siderophore-related genes were upregulated in the mutant strain in iron scarcity. Concomitantly, S. clavuligerus Δ3199 produced 1.65 and 1.9 times more catechol and hydroxamate-type siderophores, respectively, than that of the wild type strain under iron depletion. Iron containing chemically defined medium did not favor antibiotic production in S. clavuligerus Δ3199 while fermentation in starch-asparagine medium led to improved cephamycin C (2.23-fold) and clavulanic acid (2.56-fold) production in the mutant compared to the control. However, better tunicamycin yield (2.64-fold) was obtained in trypticase soy broth-grown cultures of S. clavuligerus Δ3199. Our findings demonstrate that the SCLAV_3199 gene plays a significant role in regulating both iron homeostasis and secondary metabolite biosynthesis in S. clavuligerus.
Fungal bioremediation is a very attractive tool to cope with environmental pollution. We aimed to decipher the cadmium (Cd) response of
Purpureocillium
sp. CB1, isolated from polluted soil, at ...transcriptome level by RNA-sequencing (RNA-seq). We used 500 and 2500 mg/L of Cd
2+
concentrations at two time points (t
6;36
). RNA-seq determined 620 genes that were co-expressed in all samples. The highest number of differentially expressed genes (DEGs) was obtained within the first six h of exposure to 2500 mg/L of Cd
2+
. Several genes encoding transcriptional regulators, transporters, heat shock proteins, and oxidative stress-related genes were differentially expressed under Cd
2+
stress. Remarkably, the genes that encode salicylate hydroxylase, which is involved in naphthalene biodegradation pathway, were significantly overexpressed. Utilization of diesel as the sole carbon source by CB1 even in the presence of Cd
2+
supported concomitant upregulation of hydrocarbon degradation pathway genes. Furthermore, leucinostatin-related gene expression levels increased under Cd
2+
stress. In addition, leucinostatin extracts from Cd
2+
-treated CB1 cultures showed higher antifungal activity than the control. Notably, Cd
2+
in CB1 was mainly found as bound to the cell wall, thus confirming its adsorption potential. Cd
2+
stress slightly reduced growth and led to mycelial malformation due to Cd
2+
adsorption, especially at a concentration of 2500 mg/L at t
36
. A strong correlation was recorded between RNA-seq and reverse-transcriptase-quantitative polymerase chain reaction (RT-qPCR) data. In conclusion, the study represents the first transcriptome analysis of
Purpureocillium
sp. under Cd
2+
stress, providing insights into the primary targets for rational engineering to construct strains with remarkable bioremediation potency.
Key points
• Upregulation of genes encoding salicylate hydroxylases under Cd
2+
stress
• Maximum Cd
2+
adsorption at 500 mg/L at t
36
as tightly bound to the cell wall
• Concordant bioremediation potential of CB1 on Cd
2+
and diesel
Graphical abstract
lysA
gene encoding meso-diaminopimelic acid (DAP) decarboxylase enzyme that catalyzes
l
-lysine biosynthesis in the aspartate pathway in
Streptomyces clavuligerus
was overexpressed, and its effects ...on cephamycin C (CephC), clavulanic acid (CA), and tunicamycin productions were investigated. Multicopy expression of
lysA
gene under the control of
glpF
promoter (
glpFp
) in
S
.
clavuligerus
pCOlysA led to higher expression levels ranging from 2- to 6-fold increase at both
lysA
gene and CephC biosynthetic gene cluster at T
36
and T
48
of TSBG fermentation. These results accorded well with CephC production. Thus, 1.86- and 3.14-fold higher volumetric as well as 1.26- and 1.71-fold increased specific CephC yields were recorded in
S
.
clavuligerus
pCOlysA in comparison with the wild-type and its control strain, respectively, at 48th h. Increasing the expression of
lysA
provided 4.3 times more tunicamycin yields in the recombinant strain. These findings suggested that
lysA
overexpression in
S
.
clavuligerus
made the strain more productive for CephC and tunicamycin. The results also supported the presence of complex interactions among antibiotic biosynthesis pathways in
S
.
clavuligerus
.
Polyhydroxyalkanoates (PHAs) are leading “green” alternatives for the production of biodegradable plastics. They accumulate inside archaea and bacteria as the sources of carbon and energy under ...stress conditions. This study evaluates the biosynthesis of poly(3-hydroxybutyrate) (PHB), one of the major PHAs, in the extremely halophilic archaeon
Haloarcula
sp. TG1 by microscopic, spectroscopic, and thermal analyses. The PHB production by TG1 strain was optimized in terms of growth conditions and media. The optimum PHB content (34.6% of the biomass) was obtained in Mineral Salt Medium (MSM) cultures containing 5 M NaCl and 4% glucose, at pH 7.35, incubated for 72 h at 37 °C. Replacement of glucose with starch or glycerol lowered the PHB content in biomass. Alternatively, agricultural wastes (sugar beet pulp, corn cob, and hazelnut husk) were used as cheap carbon sources. The highest PHB content (45.6% of the biomass) was obtained using sugar beet pulp treated with recombinant endoglucanase (rCKT3eng), while PHB amount was 17.8% of the biomass with chemically hydrolyzed sugar beet pulp. In conclusion, PHB production by
Haloarcula
sp. TG1 was shown to be promising biotechnologically, using a low-cost fermentation medium at optimal culture conditions.
Halophilic cellulases are indispensable enzymes of heavy industrial processes as resistant biocatalysts due to high level activity at extreme conditions. In this study, crude cellulase from an ...extreme halophilic Haloarcula sp. CKT3 was characterized. Then, recombinant expression of putative endo-1,4-β-glucanase gene, of CKT3 strain, in E. coli BL21(DE3) was performed with the aim of obtaining highly pure, active and robust industrial enzyme for such industrial aplications. The crude cellulase had optimal activity (16.9 U/mg) at 70 °C, pH 7.0 and 4 M NaCl exhibiting good thermostability, high pH and halotolerance. Indeed, it is very stable in water-insoluble organic solvents with log Po/w ≥ 2.13 and highly resistant to SDS (10%). Recombinant CKT3eng has a molecular weight of 36.9 kDa and 99% aminoacid identity to endo-l,4-β-D-glucanase from Haloarcula argentinensis. Its 3D structure was predicted using Phyre2 and I-TASSER. rCKT3eng enzyme provided 31.6 U/mg activity at optimal 50 °C, pH 7.0 and 3 M NaCl. In addition to its quite similar stability values and resistance to organic solvents and SDS, rCKT3eng has superiority over crude enzyme with 1.87-fold higher specific activity. Therefore, rCKT3eng offers a promising enzyme for industrial use with its valuable activity and stability in extreme conditions.
Indole-based heterocyclic compounds play important roles in pharmaceutical chemistry due to their unexpected biological and pharmacological properties.
Herein, we describe novel biological properties ...(antioxidant, antimicrobial and anti-cancer) of 3- bromo-1-ethyl-1H-indole (BEI) structure.
BEI was synthesized from 1-Methyl-2-phenylindole and N-bromosuccinimide and was characterized by using 1H and 13C NMR. Cytotoxicity was determined by MTT assay. Apoptosis analysis of BEI was determined by Arthur™ image-based Cytometer. Different methods were applied to assess the antioxidant activity of BEI. Molecular docking studies were conducted to determine the interactions of bonding between GST isozymes and BEI.
According to the antioxidant and antimicrobial activity assays, BEI compound showed reduced total antioxidant activity compared to the Trolox standard, whereas it showed moderate antimicrobial activity against Aspergillus niger and Phytophora eryhtrospora. Notably, the BEI compound demonstrated substantial selective cytotoxicity for the first time towards cancer cell lines, and there existed a significant decrease in the percentage of live cells treated with BEI, in comparison to the control ones. Interestingly, BEI exhibited a promising glutathione S-transferase isozymes inhibition.
The results of this study suggest that BEI seems to be a promising molecule to be used in the design of new anti-cancer agents that provide superiority to present commercial anti-cancer drugs.
Streptomyces clavuligerus is prolific producer of cephamycin C, a medically important antibiotic. In our former study, cephamycin C titer was 2-fold improved by disrupting homoserine dehydrogenase ...(hom) gene of aspartate pahway in Streptomyces clavuligerus NRRL 3585.
In this article, we aimed to provide a comprehensive understanding at the proteome level on potential complex metabolic changes as a consequence of hom disruption in Streptomyces clavuligerus AK39.
A comparative proteomics study was carried out between the wild type and its hom disrupted AK39 strain by 2 Dimensional Electrophoresis-Matrix Assisted Laser Desorption and Ionization Time-Of-Flight Mass Spectrometry (2DE MALDI-TOF/MS) and Nanoscale Liquid Chromatography- Tandem Mass Spectrometry (nanoLC-MS/MS) analyses. Clusters of Orthologous Groups (COG) database was used to determine the functional categories of the proteins. The theoretical pI and Mw values of the proteins were calculated using Expasy pI/Mw tool.
"Hypothetical/Unknown" and "Secondary Metabolism" were the most prominent categories of the differentially expressed proteins. Upto 8.7-fold increased level of the positive regulator CcaR was a key finding since CcaR was shown to bind to cefF promoter thereby direcly controlling its expression. Consistently, CeaS2, the first enzyme of CA biosynthetic pathway, was 3.3- fold elevated. There were also many underrepresented proteins associated with the biosynthesis of several Non-Ribosomal Peptide Synthases (NRPSs), clavams, hybrid NRPS/Polyketide synthases (PKSs) and tunicamycin. The most conspicuously underrepresented protein of amino acid metabolism was 4-Hydroxyphenylpyruvate dioxygenase (HppD) acting in tyrosine catabolism. The levels of a Two Component System (TCS) response regulator containing a CheY-like receiver domain and an HTH DNA-binding domain as well as DNA-binding protein HU were elevated while a TetR-family transcriptional regulator was underexpressed.
The results obtained herein will aid in finding out new targets for further improvement of cephamycin C production in Streptomyces clavuligerus.
PDF
