Presence Of Inducer Research Articles

Stimulation of mouse hematopoietic stem cells by angiogenin and DNA preparations.

Immature hematopoietic progenitors are a constant source for renewal of hemocyte populations and the basic component of the tissue and cell repair apparatus. A unique property of these cells of internalizing extracellular double-stranded DNA has been previously shown. The leukostimulatory effect demonstrated in our pioneering studies was considered to be due to the feature of this cell. In the present research, we have analyzed the effects of DNA genome reconstructor preparation (DNAgr), DNAmix, and human recombinant angiogenin on both hematopoietic stem cells and multipotent progenitors. Treatment with bone marrow cells of experimental mice with these preparations stimulates colony formation by hematopoietic stem cells and proliferation of multipotent descendants. The main lineage responsible for this is the granulocyte-macrophage hematopoietic lineage. Using fluorescent microscopy as well as FACS assay, co-localization of primitive c-Kit- and Sca-1-positive progenitors and the TAMRA-labeled double-stranded DNA has been shown. Human recombinant angiogenin was used as a reference agent. Cells with specific markers were quantified in intact bone marrow and colonies grown in the presence of inducers. Quantitative analysis revealed that a total of 14,000 fragment copies of 500 bp, which is 0.2% of the haploid genome, can be delivered into early progenitors. Extracellular double-stranded DNA fragments stimulated the colony formation in early hematopoietic progenitors from the bone marrow, which assumed their effect on cells in G0. The observed number of Sca1+/c-Kit+ cells in colonies testifies to the possibility of both symmetrical and asymmetrical division of the initial hematopoietic stem cell and its progeny.

Nonsteroidal anti-inflammatory drugs as antipyretics and modulators of a molecular clock(s) in the appendix of Sauromatum venosum inflorescence.

The appendix of the Sauromatum senosum inflorescence is a striking example of thermogenesis in plants. On the day of opening, the Sauromatum appendix becomes hot, reaching up to 32 °C. Aspirin, salicylic acid and 2,6-dihydroxybenzoic acid, a subclass of NSAIDs, induce a temperature rise from three mitochondrial sources: alternative oxidase, F1 FO -ATP synthase and adenine nucleotide translocator. This temperature rise is synchronized and compounded under various light/dark regimes. We studied the effect of different subgroups of NSAIDs on the temperature rise. Tissue slices of appendix of Sauromatum and Arum italicum inflorescences at a pre-mature stage were treated with the three inducers in combination with one NSAID under constant light or darkness and under different photoperiods. Temperature rise generated by the three heat sources in the presence of inducers and different non-selective NSAIDs were not compounded and occurred at three different times. Under constant light, DuP-697, ibuprofen, flurbiprofen, acetaminophen and diclofenac suppressed the temperature rise induced by the three salicylates. Desynchronization and delayed temperature rise were detected with 6/42-h light/ dark and 15/33-h light/dark regimes in the presence of celecoxib and ibuprofen. With a 24/24-h light/dark regime, temperature rise was suppressed in the presence of ibuprofen. There were differences in response to individual NSAIDs between appendix tissue of A. italicum and S. venosum. Mitochondrial energy balance is affected by NSAIDs. There is an interaction between light/dark regime and temperature rise and a relationship between timing mechanism and temperature rise.

PCB-77 biodegradation potential of biosurfactant producing bacterial isolates recovered from contaminated soil.

Polychlorinated biphenyls (PCBs) are persistent organic pollutants widely distributed in the environment and possess deleterious health effects. The main objective of the study was to obtain bacterial isolates from PCB-contaminated soil for enhanced biodegradation of PCB-77. Selective enrichment resulted in the isolation of 33 strains of PCB-contaminated soil nearby Bhilai steel plant, Chhattisgarh, India. Based on the prominent growth using biphenyl as the sole carbon source and the confirmation of its degradation by GC-MS/MS analysis, four isolates were selected for further study. The isolates identified by 16S rRNA gene sequencing were Pseudomonas aeruginosa MAPB-2, Pseudomonas plecoglossicida MAPB-6, Brucella anthropi MAPB-9, and Priestia megaterium MAPB-27. The isolate MAPB-9 showed a degradation of 66.15% biphenyl, while MAPB-2, MAPB-6, and MAPB-27 showed a degradation of 62.06, 57.02, and 56.55%, respectively in 48 h. Additionally, the degradation ability of these strains was enhanced with addition of co-metabolite glucose (0.2%) in the culture medium. Addition of glucose showed 100% degradation of biphenyl by MAPB-9, in 48 h, while MAPB-6, MAPB-2, and MAPB-27 showed 97.1, 67.5, and 53.3% degradation, respectively as analyzed by GC-MS/MS. Furthermore, in the presence of inducer, PCB-77 was found to be 59.89, 30.49, 27.19, and 4.43% degraded by MAPB-6, MAPB-9, MAPB-2, and MAPB-27, respectively in 7 d. The production of biosurfactants that aid in biodegradation process were observed in all the isolates. This was confirmed by ATR-FTIR analysis that showed the presence of major functional groups (CH2, CH3, CH, = CH2, C–O–C, C-O) of the biosurfactant. The biosurfactants were further identified by HPTLC and GC-MS/MS analysis. Present study is the first to report PCB-77 degradation potential of Pseudomonas aeruginosa, B. anthropi, Pseudomonas plecoglossicida, and Priestia megaterium. Similarly, this is the first report on Pseudomonas plecoglossicida and Priestia megaterium for PCB biodegradation. Our results suggest that the above isolates can be used for the biodegradation of biphenyl and PCB-77 in PCB-contaminated soil.

Major cellular and molecular processes and clinical outcomes in bone regeneration for successful dental implantation: a systematic review

Introduction: In the context of bone regeneration, bone defects still represent a major challenge in oral and maxillofacial clinical and surgical treatment. The biomimetic design of biomaterials by simulating the natural structure and composition of bone tissue has gradually become a point of research interest due to its advantages of simplicity and efficiency. Objective: It was to carry out a systematic review on cellular and molecular processes, as well as to present the main clinical approaches of bone regeneration for dental implants. Methods: The present study followed a systematic review model, following the rules of systematic review – PRISMA. The search strategy was performed in the PubMed, Cochrane Library, Web of Science and Scopus, and Google Scholar databases. The quality of the studies was based on the GRADE instrument and the risk of bias was analyzed according to the Cochrane instrument. Results: A total of 142 articles were found. A total of 84 articles were fully evaluated and 33 were included in this study. Literary findings have shown that the lack of bone in the alveolar ridges has been a major problem in functional aesthetic recovery in patients who have suffered dentoalveolar trauma. The osteoinduction process is influenced by several factors, requiring the presence of inducers, which include β-glycerolphosphate, ascorbic acid, and dexamethasone. Mesenchymal stem cells acquire the morphology and components of osteoblastic membranes and begin to express alkaline phosphatase to deposit extracellular matrix rich in calcium and certain proteins, such as osteopontin and osteocalcin. The Bio Oss® (Geistlich) biomaterial, because it is biodegradable, biocompatible, non-toxic, and has low immunogenicity and bio stimulators, can act in the regeneration of bone tissue. Application of FRP and implant placement provides stable clinical results for severely atrophic maxilla 2-4 mm. Bovine xenograft alone and in combination with liquid FRP are both successful in achieving bone augmentation around implants and produce a small change in marginal bone level and a high implant survival rate after loading. Conclusion: Through the results of the present study, it was evidenced that the success of the dental implant is directly related to successful osseointegration.

Major approaches to bucomaxillofacial bone regeneration and remodeling with the use of biomaterials: a systematic review

Introduction: The number of dental implants in the world has increased, totaling more than one million procedures per year. The development of biomaterials has represented an important therapeutic tool in the correction of bone defects. The bone regeneration process is initiated by successive mitosis of mesenchymal and endothelial stem cells, as well as by activation of osteoblasts and vascular proliferation guided by platelet-derived growth factors (PDGF) and TGF-β. Objective: To present, through a systematic review, the main results involving bone formation and remodeling in the maxillofacial system using biomaterials, cells and molecules in the appropriate biological niche of human bone. Methods: The research was carried out from July 2021 to August 2021 and developed based on Scopus, PubMed, Science Direct, Scielo, and Google Scholar, following the Systematic Review-PRISMA rules. The quality of the studies was based on the GRADE instrument and the risk of bias was analyzed according to the Cochrane instrument. Results: A total of 225 articles were found. After the selection process, 95 articles were recruited for analysis, 44 articles were selected and 43 articles were used in this study to compose the textual part. The selected articles had moderate quality in their clinical trials. The bias risks found do not affect the reliability of the results. As main findings, it was found that the osteoinduction process is influenced by several factors, requiring the presence of inducers, which include β-glycerolphosphate, ascorbic acid and dexamethasone. Mesenchymal cells acquire the morphology and components of osteoblastic membranes and start to express alkaline phosphatase to deposit extracellular matrix rich in calcium and certain proteins, such as osteopontin and osteocalcin. Bone morphogenetic proteins function as growth factors with a specific role in the proliferation and differentiation of mesenchymal stem cells present in the lesion's niche. Platelet-rich fibrin (PRF) stimulates bone regeneration more efficiently. Studies have reported that the addition of PRF with bone graft is associated with positive clinical results and is a good method of manipulating the bone graft during insertion into the maxillary sinuses and it stimulates bone regeneration. Conclusion: Optimized bone regeneration is a matter of great research to accelerate the osseointegration process, leading to reduced waiting time before any subsequent procedure. The balance between the biomaterial, mesenchymal stem cells, fibrin formation and platelet activation are responsible for the process and performance of bone regeneration or formation.

Influence of carbon, nitrogen sources, inducers, and substrates on lignocellulolytic enzyme activities of Morchella spongiola

The influence of various carbon, nitrogen sources, inducers, and different agro-wastes on the growth and ligninolytic enzyme activities of Morchella spongiola (MR17) was studied. This fungus has grown well in mineral salts broth containing mannitol as the carbon and yeast extract as the nitrogen sources. In presence of inducers, the growth was inhibited compared to control. The carbon and nitrogen sources and the inducers affected the laccase activity of M. spongiola . Rhamnose and galactose supported the laccase activity more than other carbon sources tested. The manganese peroxidase (MnP) and versatile peroxidase (VP) activities were not detected in the medium amended with different carbon and nitrogen sources. The inducers, ABTS, and veratryl alcohol enhanced the laccase activity more than others. The MnP activity was maximum in wheat straw, followed by rice straw and phenol red. Maximum VP activity was observed in wheat straw, and rice straw, followed by veratryl alcohol. The protein content increased significantly in groundnuts shell and wheat grain substrates inoculated with M. spongiola . Wheat grains and wheat bran served as the most promising substrates with maximum lignocellulolytic activity. The laccase activity was higher in wheat grains and wheat bran, while VP and MnP activities were higher in rice straw and wheat straw. The cellulase activity was much less compared to other enzyme activities in all the substrates. Maximum lignin degradation was observed in wheat straw, followed by wheat bran. The cellulose degradation was significantly higher in groundnuts shell compared to other substrates. The present study results suggest that M. spongiola can utilize these substrates, induce enzyme activities, and have the potential for mushroom cultivation.

Role of endopeptidases in peptidoglycan synthesis mediated by alternative cross-linking enzymes in Escherichia coli.

Bacteria resist to the turgor pressure of the cytoplasm through a net-like macromolecule, the peptidoglycan, made of glycan strands connected via peptides cross-linked by penicillin-binding proteins (PBPs). We recently reported the emergence of β-lactam resistance resulting from a bypass of PBPs by the YcbB L,D-transpeptidase (LdtD), which form chemically distinct 3→3 cross-links compared to 4→3 formed by PBPs. Here we show that peptidoglycan expansion requires controlled hydrolysis of cross-links and identify among eight endopeptidase paralogues the minimum enzyme complements essential for bacterial growth with 4→3 (MepM) and 3→3 (MepM and MepK) cross-links. Purified Mep endopeptidases unexpectedly displayed a 4→3 and 3→3 dual specificity implying recognition of a common motif in the two cross-link types. Uncoupling of the polymerization of glycan chains from the 4→3 cross-linking reaction was found to facilitate the bypass of PBPs by YcbB. These results illustrate the plasticity of the peptidoglycan polymerization machinery in response to the selective pressure of β-lactams.

Overexpression and repression of key rate-limiting enzymes (acetyl CoA carboxylase and HMG reductase) to enhance fatty acid production from Rhodotorula mucilaginosa.

Implementing two-way strategies to enhance the lipid production in Rhodotorula mucilaginosa with the help of metabolic engineering was focused on the overexpression of acetyl coenzyme A carboxylase (ACC1 carboxylase) gene and repression of 3-hydroxy 3-methylglutaryl reductase (HMG-CoA reductase). Using an inducer (sodium citrate) and inhibitor (rosuvastatin), the amounts of biomass, lipid, and carotenoid were estimated. In the presence of inhibitor (200 mM), 62% higher lipid concentration was observed, while 44% enhancement was recorded when inducer (3 mM) was used. A combination of both inhibitor and inducer resulted in a 57% increase in lipid concentration by the oleaginous yeast. These results were again confirmed by real-time polymerase chain reaction by targeting the expression of the genes coding for ACC1 carboxylase and 13-fold increase was recorded in the presence of inducer as compared with control. This combined strategy (inducer and inhibitor use) has been reported for the first time as far as the best of our knowledge. The metabolic engineering strategies reported here will be a powerful approach for the enhanced commercial production of lipids.

Aerobic degradation of cis-dichloroethene by the marine bacterium Marinobacter salsuginis strain 5N-3.

An aerobic bacterium, designated strain 5N-3 (NBRC 113055), that degrades cis-dichloroethene (cDCE) was isolated from a sea sediment in Japan. Strain 5N-3 was able to degrade a certain amount of cDCE in the presence of pyruvate without the action of inducers. In the presence of inducers, such as phenol and benzene, the strain completely removed cDCE. By the application of 16S ribosomal RNA (16S rRNA) gene sequencing and average nucleotide identity analyses, the strain 5N-3 was identified as Marinobacter salsuginis. On the other hand, identified species of Marinobacter are not known to degrade cDCE at all. A draft genome sequence analysis of the strain 5N-3 suggested that the dmp-homologous operon (operon for phenol degradation) may be contributing to the aerobic degradation of cDCE. This is the first report on an aerobic marine bacterium that has been found to degrade cDCE.

The Interaction between Marasmius pulcheripes J8 and Soil Fungi on Laccase Activity for POME Degradation

A study on the effect of Marasmius pulcheripes J8 and soil fungi interaction on the activity and ability of laccase to degrade palm oil mill effluent (POME) was previously conducted, and some Basidiomycetes fungi were identified capable. Therefore, the aim of this study was to determine the ability of Marasmius pulcheripes J8 to degrade POME in the presence of inducers , and interactions with soil fungi. Furthermore, 3 types of inducers were applied to elevate its laccase activity, which include CuSO 4 , sucrose and Ammonium tartrate. In addition, M. pulcheripes J8 was grown together with soil fungi, encompassing Aspergilus niger NK and Penicillium sp R 75, in order to boost the action . T he results showed the highest laccase activity was in M. pulcheripes J8 pure culture on a PDB medium of 3566.04 U / mL. Moreover, the POME decolorization was up to 74.25% after 20 days of incubation, and reduced COD level was 81%. Meanwhile, the addition of an inducer has never been performed before, and the outcome of this investigation showed the ability of M. pulcheripes J8 to degrade POME, and decrease environmental pollution. POME waste treatment using fungi is more affordable than other methods.

ISOENZIMAS Y ACTIVIDAD DE LACASAS PRODUCIDAS POR Pleurotus ostreatus CRECIDO A DIFERENTES TEMPERATURAS

Pleurotus ostreatus produces laccase isoforms depending on the conditions in which it grows, it has been suggested that the presence of inducers, the pH of the culture medium as well as the type and concentration of carbon and nitrogen sources promote the production of these isoforms, however, the effect of temperature of Pleurotus ostreatus growth on its production of laccase isoforms has not been evaluated. In this study, Pleurotus ostreatus was grown in solid-state fermentation at different temperatures (10, 15, 20, 25, 30 and 35 °C) and the activity and isoforms number of laccases produced in each bioprocess was determined. At 10 and 35 °C there was no growth of the fungus. The activity of laccases was different in each fermentation, being at 25 ºC where the highest value was observed (20000 U L-1). The laccase isoenzymes pattern was also different depending on the incubation temperature of the fungus, it was observed in some times of the culture, 4 isoforms in the fermentations incubated at 15 and 20 ºC, 3 isoforms at 25 ºC and 2 isoforms at 30 ºC. These results show that the growth temperature of Pleurotus ostreatus modifies its patterns of activity and of isoforms of laccases.

Characterisation of the bacterial biosensor GMG in E‐coli BL21 (DE3)

In this work, a gold biosensor has been constructed and characterised in Escherichia coli (E. coli). Though gold biosensors have been previously reported, a NOT gate gold biosensing genetic circuit was constructed using an orthogonal inverter system as an alternative to a circuit where output is contingent solely on input. This enables the detection of changes in gold concentration whereby circuit remains OFF in presence of inducer and ON in absence of inducer. The proposed biosensor is an alternative to conventional forms of gold detection such as Inductively Coupled Plasma-Mass Spectrometry (ICP-MS). The constructed biosensor is a collection of components – golTSB operon (isolated from Salmonella enterica), mf-Lon protease (isolated from Mesoplasma florum) and associated ssrA degradation tag with GFP acting as a reporter. The minimum time and concentration required by the biosensor gol-mflon-GFP (GMG) to achieve the ON output is characterised here using plate reader data derived from a single multi-stage assay.

Mutations in AraR leading to constitutive of arabinolytic genes in Aspergillus niger under derepressing conditions

The AraR transcription factor of Aspergillus niger encodes a Zn(II)2Cys6 transcription factor required for the induction of genes encoding arabinolytic enzymes. One of the target genes of AraR is abfA, encoding an arabinofuranosidase. The expression of abfA as well as other L-arabinose-induced genes in A. niger requires the presence of L-arabinose or its derivative L-arabitol as an inducer to activate AraR-dependant gene expression. In this study, mutants were isolated that express L-arabinose-induced genes independently of the presence of an inducer under derepressing conditions. To obtain these mutants, a reporter strain was constructed in a ΔcreA background containing the L-arabinose-responsive promoter (PabfA) fused to the acetamidase (amdS) gene. Spores of the ΔcreA PabfA-amdS reporter strain were UV-mutagenized and mutants were obtained by their ability to grow on acetamide without the presence of inducer. From a total of 164 mutants, 15 mutants were identified to contain transacting mutations resulting in high arabinofuranosidase activity in the medium after growth under non-inducing conditions. Sequencing of the araR gene of the 15 constitutive mutants revealed that 14 mutants carried a mutation in AraR. Some mutations were found more than once and in total nine different point mutations were identified in AraR. The AraRN806I point mutation was reintroduced into a parental strain and confirmed that this point mutation leads to inducer-independent expression of AraR target genes. The inducer independent of L-arabinose-induced genes in the AraRN806I mutant was found to be sensitive to carbon catabolite repression, indicating that the CreA-mediated carbon catabolite repression is dominant over the AraRN806I mutant allele. These mutations in AraR provide new opportunities to improve arabinase production in industrial fungal strains.

Conidial germination of marine-derived fungi under simulated deep-sea conditions in the presence of inducers

The abilities of conidia of four deep-sea fungi, Aspergillus flavus NIOCC20, A. sydowii CH2, A. terreus A4636 and Penicillium sp. A4623 and one terrestrial fungus, A. terreus MTCC479 to germinate under simulated deep-sea conditions were investigated by exposing the conidia to low temperature (4 °C) and elevated hydrostatic pressures (10, 20, 30, 40, 50 and 60 MPa). The conidial germination was evaluated in presence of chemical inducers, including Thiamine, Sodium pyruvate, Hydrochloric acid, Sulphuric acid and Sucrose, in a basal medium of 1/5th strength of malt extract broth and subjected to heat treatment at 70 °C for 60 min, and incubated for 30 days under simulated deep-sea conditions. The conidia of Aspergillus flavus, A. sydowii and Penicillium sp. germinated and developed into mycelia under almost all the treatments with and without heat inducement. However, the number of germinated conidia of A. terreus A4636 and A. terreus MTCC479 was lower than the other three fungal species. This is the first study demonstrating in vitro germination of conidia of deep-sea fungi at low temperature with elevated hydrostatic pressures. Further studies are warranted to obtain insights into in situ conditions that are favourable for conidial germination and fungal activity in deep-sea niches.

The Control of Apoptotic Death in the Cells of Granulomatous Inflammatory Lesions from Mice with Latent Tuberculous Infection in the Ex Vivo Model

Tuberculosis is a leading worldwide health problem. The latent, symptom-free stage of tuberculous infection is characterized by the formation of granulomas, specific aggregates of immune cells, predominantly macrophages, containing mycobacteria. The apoptotic death of macrophages containing mycobacteria is considered the main mechanism by which animals and human organisms oppose tuberculous infection and control its development. Previously, we have comparedMycobacterium-host cell relationships in individual granuloma cells from mice with latent tuberculous infection and cells from mouse bone marrow and peritoneal cultures infected with BCG vaccine in vitro and shown that increased death rates were revealed for macrophages heavily loaded with mycobacteria after acute BCG infection in vitro. While in ex vivo cultures granuloma macrophages with large numbers of BCG mycobacteria in them were still viable and had neither apoptotic nor necrotic morphology. Since different specific cellular responses to latent chronic and acute BCG infection in mouse cells were determined, the our aim was to analyze granulomas isolated from the lungs, spleens and bone marrow of Balb/c mice with latent BCG infection for the presence of inducers and markers of apoptotic cell death. In granuloma cells with increased levels of the inducer of apoptosis TNFα, proapoptotic proteins Вах and Ваd, death receptor Fas/ CD95 and scavenge receptor CD36, we did not observe P53 stabilization or caspase-3 activation in the cytoplasm or nuclei of macrophages and dendritic cells, irrespective of the presence or absence of acid-fast BCG mycobacteria in them. The survival receptor CD30 was detected on the cell membranes of only few granuloma macrophages. However, at later times of tuberculous infection in mice, virtually all macrophages and other granuloma cell types had considerable amounts of the antiapoptotic protein Bcl-2 in the cytoplasm and, probably, mitochondria, in contrast to macrophages from bone barrow cell cultures and peritoneal exudates infected with BCG mycobacteria in vitro. Preservation of mitochondrial ΔΨm during staining of living granuloma macrophages containing large amounts of the Bcl-2 protein was indicative of its involvement in maintaining the integrity of mitochondrial elements and the protection of granuloma cells from death, because in similar experiments the control macrophages that did not have any Bcl-2 protein in them had considerably reduced ΔΨm and exhibited morphological signs of apoptotic death. Taken together, our results suggest that the antiapoptotic protein Bcl-2 has been proposed to contribute to the viability of granulomas macrophages not only in ex vivo culture, but also in the animal organism when faced with mycobacterial, proinflammatory and proapoptotic factors operating in granulomatous inflammatory lesions at various times of latent tuberculous infection in mice.

Improvement of the reverse tetracycline transactivator by single amino acid substitutions that reduce leaky target gene expression to undetectable levels.

Conditional gene expression systems that enable inducible and reversible transcriptional control are essential research tools and have broad applications in biomedicine and biotechnology. The reverse tetracycline transcriptional activator is a canonical system for engineered gene expression control that enables graded and gratuitous modulation of target gene transcription in eukaryotes from yeast to human cell lines and transgenic animals. However, the system has a tendency to activate transcription even in the absence of tetracycline and this leaky target gene expression impedes its use. Here, we identify single amino-acid substitutions that greatly enhance the dynamic range of the system in yeast by reducing leaky transcription to undetectable levels while retaining high expression capacity in the presence of inducer. While the mutations increase the inducer concentration required for full induction, additional sensitivity-enhancing mutations can compensate for this effect and confer a high degree of robustness to the system. The novel transactivator variants will be useful in applications where tight and tunable regulation of gene expression is paramount.

English

Laccases are enzymes that have a great potential for use in breaking down toxic compounds. Fungal laccases show high enzymatic activity, especially those produced by basidiomycetes. Depending on the culture conditions and the strain used, a variety of isoenzymes and/or enzymatic activities can be obtained. In this study, extracellular laccase enzymes produced by Pleurotus ostreatus was identified in a submerged culture (SmF), with and without copper sulphate as a chemical inducer, and in a solid-state culture (SSF), using wheat straw as natural inducer. This study was conducted to observe the expression of the enzymes produced under the culture conditions tested and their persistence during the culture, as well as the extracellular activity produced and the correspondence that the isoenzymes presented between the intracellular and extracellular media. A positive effect of the inducers on the specific laccase activity was observed either in SmF with copper sulphate or SSF (41.11 and 40.43 UI/mg protein, respectively), compared with that obtained in SmF without copper sulphate (2.87 UI/mg protein). This effect was different only at the time when the highest activity appeared (360 and 120 h, respectively), showing advantages in SSF. The same three isoenzymes were observed in the three kinds of cultures. The main differences among the laccase profiles reside in the time when they appeared in each culture and only an additional form of lower molecular weight was observed in SSF. The laccase enzymes in the intracellular extracts were equal to those in the extracellular ones. The laccase isoenzymes profiles suggest that the presence of inducers helps in maintaining the activity through the culture time. Key words: Phenol oxidases, basidiomycete, enzymatic activity, copper, wheat straw, solid-state culture (SSF), submerged culture (SMF).&nbsp

Simultaneous Real-Time Monitoring of Oxygen Consumption and Hydrogen Peroxide Production in Cells Using Our Newly Developed Chip-Type Biosensor Device

All living organisms bear its defense mechanism. Immune cells during invasion by foreign body undergoes phagocytosis during which monocyte and neutrophil produces reactive oxygen species (ROS). The ROS generated in animal cells are known to be involved in several diseases and ailments, when generated in excess. Therefore, if the ROS generated in cells can be measured and analyzed precisely, it can be employed in immune function evaluation and disease detection. The aim of the current study is to introduce our newly developed chip-type biosensor device with high specificity and sensitivity. It comprises of counter electrode and working electrodes I and II. The counter electrode is a platinum plate while the working electrodes I and II are platinum microelectrode and osmium-horseradish peroxidase modified gold electrode, respectively which acts as oxygen and hydrogen peroxide (H2O2) detection sensors. Simultaneous measurement of oxygen consumption and H2O2 generation were measured in animal cells under the effect of exogenous addition of differentiation inducer, phorbol 12-myristate 13-acetate. The results obtained showed considerable changes in reduction currents in the absence and presence of inducer. Our newly developed chip-type biosensor device is claimed to be a useful tool for real-time monitoring of the respiratory activity and precise detection of H2O2 in cells. It can thus be widely applied in biomedical research and in clinical trials being an advancement over other H2O2 detection techniques.

Molecular level biodegradation of phenol and its derivatives through dmp operon of Pseudomonas putida: A bio-molecular modeling and docking analysis

Participation of Pseudomonas putida-derived methyl phenol (dmp) operon and DmpR protein in the biodegradation of phenol or other harmful, organic, toxic pollutants was investigated at a molecular level. Documentation documents that P. putida has DmpR protein which positively regulates dmp operon in the presence of inducers; like phenols. From the operon, phenol hydroxylase encoded by dmpN gene, participates in degrading phenols after dmp operon is expressed. For the purpose, the 3-D models of the four domains from DmpR protein and of the DNA sequences from the two Upstream Activation Sequences (UAS) present at the promoter region of the operon were demonstrated using discrete molecular modeling techniques. The best modeled structures satisfying their stereo-chemical properties were selected in each of the cases. To stabilize the individual structures, energy optimization was performed. In the presence of inducers, probable interactions among domains and then the two independent DNA structures with the fourth domain were perused by manifold molecular docking simulations. The complex structures were made to be stable by minimizing their overall energy. Responsible amino acid residues, nucleotide bases and binding patterns for the biodegradation, were examined. In the presence of the inducers, the biodegradation process is initiated by the interaction of phe50 from the first protein domain with the inducers. Only after the interaction of the last domain with the DNA sequences individually, the operon is expressed. This novel residue level study is paramount for initiating transcription in the operon; thereby leading to expression of phenol hydroxylase followed by phenol biodegradation.

Novel phenotype of mouse spermatozoa following deletion of nine β-defensin genes.

β-defensin peptides are a large family of antimicrobial peptides. Although they kill microbes in vitro and interact with immune cells, the precise role of these genes in vivo remains uncertain. Despite their inducible presence at mucosal surfaces, their main site of expression is the epididymis. Recent evidence suggests that a major function of these peptides is in sperm maturation. In addition to previous work suggesting this, work at the MRC Human Genetics Unit, Edinburgh, has shown that homozygous deletion of a cluster of nine β-defensin genes in the mouse results in profound male sterility. The spermatozoa derived from the mutants had reduced motility and increased fragility. Epididymal spermatozoa isolated from the cauda region of the homozygous mutants demonstrated precocious capacitation and increased spontaneous acrosome reactions compared with those from wild-types. Despite this, these mutant spermatozoa had reduced ability to bind to the zona pellucida of oocytes. Ultrastructural examination revealed a disintegration of the microtubule structure of mutant-derived spermatozoa isolated from the epididymal cauda region, but not from the caput. Consistent with premature acrosome reaction and hyperactivation, spermatozoa from mutant animals had significantly increased intracellular calcium content. This work demonstrates that in vivo β-defensins are essential for successful sperm maturation, and that their disruption alters intracellular calcium levels, which most likely leads to premature activation and spontaneous acrosome reactions that result in hyperactivation and loss of microtubule structure of the axoneme. Determining which of the nine genes are responsible for the phenotype and the relevance to human sperm function is important for future work on male infertility.