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1.
Ageing Res Rev ; 94: 102171, 2024 02.
Article in English | MEDLINE | ID: mdl-38141735

ABSTRACT

Parkinson's Disease (PD) is becoming a growing global concern by being the second most prevalent disease next to Alzheimer's Disease (AD). Henceforth new exploration is needed in search of new aspects towards the disease mechanism and origin. Evidence from recent studies has clearly stated the role of Gut Microbiota (GM) in the maintenance of the brain and as a root cause of various diseases and disorders including other neurological conditions. In the case of PD, with an unknown etiology, the GM is said to have a larger impact on the disease pathophysiology. Although GM and its metabolites are crucial for maintaining the normal physiology of the host, it is an undeniable fact that there is an influence of GM in the pathophysiology of PD. As such the Enteroendocrine Cells (EECs) in the epithelium of the intestine are one of the significant regulators of the gut-brain axis and act as a communication mediator between the gut and the brain. The communication is established via the molecules of neuroendocrine which are said to have a crucial part in neurological diseases such as AD, PD, and other psychiatry-related disorders. This review is focused on understanding the proper role of GM and EECs in PD. Here, we also focus on some of the metabolites and compounds that can interact with the PD genes causing various dysfunctions in the cell and facilitating the disease conditions using bioinformatical tools. Various mechanisms concerning EECs and PD, their identification, the latest studies, and available current therapies have also been discussed.


Subject(s)
Alzheimer Disease , Gastrointestinal Microbiome , Parkinson Disease , Humans , Brain-Gut Axis , Brain
2.
Chemosphere ; 301: 134625, 2022 Aug.
Article in English | MEDLINE | ID: mdl-35439490

ABSTRACT

Parkinson's disease (PD) is a neurodegenerative condition characterized by the dopamine (DA) neuronal loss in the substantia nigra. PD impairs motor controls symptoms such as tremor, rigidity, bradykinesia and postural imbalance gradually along with non-motor problems such as olfactory dysfunction, constipation, sleeping disorder. Though surplus of factors and mechanisms have been recognized, the precise PD etiopathogenesis is not yet implied. Reports suggest that various environmental factors play a crucial role in the causality of the PD cases. Epidemiological studies have reported that heavy metals has a role in causing defects in substantia nigra region of brain in PD. Though the reason is unknown, exposure to heavy metals is reported to be an underlying factor in PD development. Metals are classified as either essential or non-essential, and they have a role in physiological processes such protein modification, electron transport, oxygen transport, redox reactions, and cell adhesion. Excessive metal levels cause oxidative stress, protein misfolding, mitochondrial malfunction, autophagy dysregulation, and apoptosis, among other things. In this review, we check out the link between heavy metals like copper (Cu), arsenic (As), cadmium (Cd), iron (Fe), and lithium (Li) in neurodegeneration, and how it impacts the pathological conditions of PD. In conclusion, increase or decrease in heavy metals involve in regulation of neuronal functions that have an impact on neurodegeneration process. Through this review, we suggest that more research is needed in this stream to bring more novel approaches for either disease modelling or therapeutics.


Subject(s)
Arsenic , Metals, Heavy , Neurotoxicity Syndromes , Parkinson Disease , Arsenic/toxicity , Cadmium , Copper , Humans , Iron/metabolism , Lithium , Metals, Heavy/toxicity
3.
Protein J ; 41(2): 327-336, 2022 04.
Article in English | MEDLINE | ID: mdl-35119603

ABSTRACT

Transgenic crops expressing Cry δ-endotoxins of Bacillus thuringiensis for insect resistance have been commercialized worldwide with increased crop productivity and spectacular socioeconomic gains. To attain the enhanced level of protein expression, the cry genes have to be extensively modified for RNA stability and translation efficiency in the plant systems. However, such modifications in nucleotide sequences make it difficult to express the cry genes in Escherichia coli because of the presence of E. coli rare codons. Induction of gene expression through the T7 promoter/lac operator system results in high levels of transcription but limits the availability of activated tRNA corresponding to rare codons that leads to translation stalling at ribosomes. In the present study, an Isopropyl ß-D-1-thiogalactopyranoside (IPTG)/rifampicin combination-based approach was adopted to induce transcription of cry genes through T7 promoter/lac operator while simultaneously inhibiting the transcription of host genes through rifampicin. The results show that the IPTG/rifampicin combination leads to high-level expression of four plant codon-optimized cry genes (cry2Aa, cry1F, cry1Ac, and cry1AcF). Northern blot analysis of the cry gene expressing E. coli samples showed that the RNA expression level in the IPTG-induced samples was higher as compared to that in the IPTG/rifampicin-induced samples. Diet overlay insect bioassay of IPTG/rifampicin-induced Cry toxins with Helicoverpa armigera larvae showed bioactivity (measured as LC50) similar to the previous studies. The experiment has proved that recombinant synthetic gene (plant codon-optimized gene) with the combination of Rifampicin which inhibits DNA-dependent bacterial RNA polymerase and reduces the excessive baggage of translational machinery of the bacterial cell triggers the production of synthetic protein. Purification of protein using high pH buffer increases the solubility of the protein. Further, LC50 analysis shows no reduction of protein activity leads to protein stability. Further, purified cry toxin protein can be used for crop protection against pests and a purified form of the synthetic protein can be used for antibody production and perform the immunoassay for the identification of the transgenic plant. The crystallographic structure of synthetic protein could be used for interaction study with another insect to see insecticidal activity.


Subject(s)
Bacillus thuringiensis , Endotoxins , Animals , Bacillus thuringiensis/genetics , Bacillus thuringiensis Toxins , Bacterial Proteins/genetics , Bacterial Proteins/pharmacology , Codon , Escherichia coli/genetics , Hemolysin Proteins/genetics , Hemolysin Proteins/pharmacology , Isopropyl Thiogalactoside , Larva , Rifampin/pharmacology
4.
Int J Biol Macromol ; 205: 185-192, 2022 Apr 30.
Article in English | MEDLINE | ID: mdl-35182560

ABSTRACT

Evolving insect resistance to delta-endotoxins can be delayed by using a few strategies like high dosage, refugia, and gene stacking which require the expression of delta-endotoxins at sufficiently high levels to kill the resistant insects. In this study, we comparatively analyzed the efficacy of targeting truncated cry1Ac protein to the cytoplasm, endoplasmic reticulum (ER), and chloroplast to obtain high protein expression. mRNA and protein profiling of cry1Ac showed that both ER and chloroplast are efficient targets for expressing high levels of truncated cry1Ac. A maximum of 0.8, 1.6, and 2.0% cry1Ac of total soluble protein were obtained when the truncated cry1Ac was expressed in the cytoplasm, routed through ER, and targeted to the chloroplast. We further showed that not only the protein content but also the biological activity of truncated cry1Ac increases by sub-cellular targeting and the biological activity is slightly greater in the ER routed transgenic lines by conducting different bioassays on Helicoverpa armigera. Using native Western analysis, we demonstrated that the truncated cry1Ac protein could exist as oligomers in plant cells and this oligomerization capability is low in the cytoplasm. In conclusion, routing of delta endotoxins through ER is the first choice to obtain high protein expression and bioactivity.


Subject(s)
Bacillus thuringiensis , Moths , Animals , Bacillus thuringiensis/genetics , Bacillus thuringiensis/metabolism , Bacillus thuringiensis Toxins , Bacterial Proteins/genetics , Bacterial Proteins/metabolism , Bacterial Proteins/pharmacology , Endotoxins/genetics , Hemolysin Proteins/genetics , Hemolysin Proteins/metabolism , Hemolysin Proteins/pharmacology , Larva/metabolism , Moths/genetics
5.
Trop Biomed ; 37(3): 560-565, 2020 Sep 01.
Article in English | MEDLINE | ID: mdl-33612771

ABSTRACT

Primary prostatic melioidosis is a rare presentation of melioidosis even in melioidosis endemic areas. We report a case of a 58-year-old man with underlying diabetes mellitus who presented with a 5-day history of high-grade fever associated with lower urinary tract symptoms. Suprapubic tenderness and tender prostatomegaly were noted on examination. An abdominal computed tomography (CT) scan confirmed the presence of a prostatic abscess. Both blood and prostatic pus cultures grew Burkholderia pseudomallei. He was initially started on intravenous ceftazidime, followed by an escalation to intravenous meropenem. He was discharged home with oral amoxicillin-clavulanate and doxycycline after completing 12 days of meropenem. Unfortunately, his compliance to oral antibiotic therapy was poor, and he succumbed to the disease.


Subject(s)
Abscess/microbiology , Anti-Bacterial Agents/therapeutic use , Burkholderia pseudomallei/isolation & purification , Medication Adherence , Melioidosis/drug therapy , Prostatic Diseases/microbiology , Fatal Outcome , Humans , Malaysia , Male , Middle Aged
6.
Tropical Biomedicine ; : 560-565, 2020.
Article in English | WPRIM (Western Pacific) | ID: wpr-825512

ABSTRACT

@#Primary prostatic melioidosis is a rare presentation of melioidosis even in melioidosis endemic areas. We report a case of a 58-year-old man with underlying diabetes mellitus who presented with a 5-day history of high-grade fever associated with lower urinary tract symptoms. Suprapubic tenderness and tender prostatomegaly were noted on examination. An abdominal computed tomography (CT) scan confirmed the presence of a prostatic abscess. Both blood and prostatic pus cultures grew Burkholderia pseudomallei. He was initially started on intravenous ceftazidime, followed by an escalation to intravenous meropenem. He was discharged home with oral amoxicillin-clavulanate and doxycycline after completing 12 days of meropenem. Unfortunately, his compliance to oral antibiotic therapy was poor, and he succumbed to the disease.

7.
Med J Armed Forces India ; 74(2): 187-189, 2018 Apr.
Article in English | MEDLINE | ID: mdl-29692491
8.
J Alzheimers Dis Rep ; 1(1): 277-286, 2017 Dec 29.
Article in English | MEDLINE | ID: mdl-30480244

ABSTRACT

DNA conformation and stability are critical for the normal cell functions, which control many cellular processes in life, such as replication, transcription, DNA repair, etc. The accumulation of amyloid-ß peptide (Aß) and Copper (Cu) are the etiological factors for neurodegenerative diseases and hypothesized that they can cause DNA instability. In the current investigation, we studied copper and Aß1-16 induced conformation and stability changes in CAG/CTG sequences and found alterations from B-DNA to altered B-conformation. Further, the interaction of the copper and Aß1-16 with CAG/CTG sequences was studied by molecular docking modeling and results indicated that the interaction of copper and Aß1-16 was through the hydrogen bond formation between adenine, guanine, and cytocine. This study illustrates the role of the copper and Aß1-16 in modulating the DNA conformation and stability.

9.
J Immunoassay Immunochem ; 37(3): 228-42, 2016.
Article in English | MEDLINE | ID: mdl-26674740

ABSTRACT

The relatively short circulatory half-life (2-3 min) of staphylokinase is a major drawback in the development of SAK- (staphylokinase) based thrombolytic drug. A rapid and sensitive method, based on indirect competitive ELISA, was developed and validated for quantitative determination of SAK in rabbit plasma. The dynamic range of the assay varied between 0.41 ± 0.16 µg/L and 9.03 ± 0.38 µg/L (R(2) = 0.98) for SAK in rabbit plasma. There were no dilution linearity issues apparent with this assay. The precision (% CV) ranged from 4.6-9.7% for the intraassay and from 17.1-19.3% for interassay. This validated method was successfully employed for evaluation of various pharmacokinetic parameters of SAK in rabbit.


Subject(s)
Enzyme-Linked Immunosorbent Assay/methods , Enzyme-Linked Immunosorbent Assay/standards , Fibrinolytic Agents/pharmacokinetics , Metalloendopeptidases/blood , Animals , Female , Fibrinolytic Agents/blood , Male , Metalloendopeptidases/metabolism , Mice , Mice, Inbred BALB C , Rabbits , Recombinant Proteins/blood , Recombinant Proteins/metabolism
10.
Appl Biochem Biotechnol ; 172(5): 2324-37, 2014 Mar.
Article in English | MEDLINE | ID: mdl-24390855

ABSTRACT

Pectinase has been an integral part of commercial food processing, where it is used for degradation of pectin and facilitates different processing steps such as liquefaction, clarification and juice extraction. The industry currently uses pectinases from mesophilic or thermophilic microorganisms which are well established, but recently, there has been is a new trend in the food industry to adopt low-temperature processing. This trend is due to the potential economic and environmental advantages which the industry envisages. In order to achieve this change, an alternative for the existing pectinases, which are mostly mesophilic and temperature-dependent, must be identified, which can function efficiently at low temperatures. Psychrophilic pectinases derived from cold-adapted microorganisms, are known to function at low to freezing temperatures and may be an alternative to address the problem. Psychrophilic pectinases can be obtained from the vast microflora inhabiting various cold regions on earth such as oceans, Polar Regions, snow-covered mountains, and glaciers. This article is intended to study the advantages of cold active pectinases, its sources, and the current state of the research.


Subject(s)
Food Handling/methods , Pectins/chemistry , Polygalacturonase/chemistry , Aspergillus niger/chemistry , Aspergillus niger/enzymology , Bacillus subtilis/chemistry , Bacillus subtilis/enzymology , Carbohydrate Sequence , Cold Climate , Cold Temperature , Erwinia/chemistry , Erwinia/enzymology , Humans , Ice Cover/microbiology , Isoenzymes/chemistry , Isoenzymes/classification , Isoenzymes/metabolism , Kinetics , Models, Molecular , Molecular Sequence Data , Polygalacturonase/classification , Polygalacturonase/metabolism
11.
Virusdisease ; 25(3): 390-3, 2014.
Article in English | MEDLINE | ID: mdl-25674610

ABSTRACT

Availability of adequate quantity of purified virus preparation from plant tissue is the major limitation in producing polyclonal antibodies (PAb) to begomovirus. Very few examples show successful utilization of E. coli expressed recombinant coat protein (CP) for immuno diagnosis of begomoviruses. In the present study, ~771 bp CP gene (~29.0 kDa) of Pumpkin yellow vein mosaic virus (PYVMV) was expressed as a ~71.0 kDa fusion protein with maltose binding protein (MBP) (~42.0 kDa) in E. coli. The MBP-CP was obtained in soluble state. The PAb to the purified fusion protein successfully detected PYVMV and other bipartite and monopartite begomoviruses in the field samples at 1:250 dilution in enzyme linked immunosorbent assay. Our study for the first time showed that MBP-tag fusion CP was suitable to produce diagnostic antibody to begomoviruses.

12.
Biomed Res Int ; 2013: 297305, 2013.
Article in English | MEDLINE | ID: mdl-23998121

ABSTRACT

The thrombolytic therapy with clinically approved drugs often ensues with recurrent thrombosis caused by thrombin-induced platelet aggregation from the clot debris. In order to minimize these problems, a staphylokinase (SAK)-based bacterial friendly multifunctional recombinant protein SRH (staphylokinase (SAK) linked with tripeptide RGD and dodecapeptide Hirulog (SRH)) was constructed to have Hirulog as an antithrombin agent and RGD (Arg-Gly-Asp) as an antiplatelet agent in the present study. This multifunctional fusion protein SRH was expressed in osmotically inducible E. coli GJ1158 as soluble form and purified with a yield of 0.27 g/L and functionally characterized in vitro. SRH retained the fibrinolytic activity and plasminogen activation rate comparable to the parental counterpart SAK. The antithrombin activity of SRH was significantly higher than SAK. The platelet rich clot lysis assay indicated that SRH had enhanced platelet binding activity and T(50%) and C50 of SRH were significantly lower than that of SAK. Furthermore, SRH inhibited the ADP-induced platelet aggregation in dose-dependent manner while SAK had no significant effect on platelet aggregation. Thus, the current study suggests that the SAK variant produced from osmotically inducible GJ1158 is more potent thrombolytic agent with antithrombin and antiplatelet aggregation activities for reduction of reocclusion in thrombolytic therapy.


Subject(s)
Escherichia coli/physiology , Metalloendopeptidases/genetics , Metalloendopeptidases/pharmacology , Platelet Aggregation/drug effects , Protein Engineering/methods , Cells, Cultured , Genetic Enhancement/methods , Genetic Variation , Humans , Metalloendopeptidases/metabolism , Platelet Aggregation Inhibitors/metabolism , Platelet Aggregation Inhibitors/pharmacology , Recombinant Fusion Proteins/biosynthesis , Recombinant Fusion Proteins/pharmacology , Salts/metabolism
13.
Prep Biochem Biotechnol ; 43(8): 766-80, 2013.
Article in English | MEDLINE | ID: mdl-23876137

ABSTRACT

In the present investigation Thalassospira frigidphilosprofundus, a novel species from the deep waters of the Bay of Bengal, was explored for the production of cold-active ß-galactosidase by submerged fermentation using marine broth medium as the basal medium. Effects of various medium constituents, namely, carbon, nitrogen source, pH, and temperature, were investigated using a conventional one-factor-at-a-time method. It was found that lactose, yeast extract, and bactopeptones are the most influential components for ß-galactosidase production. Under optimal conditions, the production of ß-galactosidase was found to be 3,864 U/mL at 20 ± 2°C, pH 6.5 ± 0.2, after 48 hr of incubation. ß-Galactosidase production was further optimized by the Taguchi orthogonal array design of experiments and the central composite rotatable design (CCRD) of response surface methodology. Under optimal experimental conditions the cold-active ß-galactosidase enzyme production from Thalassospira frigidphilosprofundus was enhanced from 3,864 U/mL to 10,657 U/mL, which is almost three times higher than the cold-active ß-galactosidase production from the well-reported psychrophile Pseudoalteromonas haloplanktis.


Subject(s)
Bacterial Proteins/biosynthesis , Bays/microbiology , Principal Component Analysis , Rhodospirillaceae/chemistry , beta-Galactosidase/biosynthesis , Bacterial Proteins/isolation & purification , Carbon/metabolism , Cold Temperature , Culture Media/chemistry , Fermentation , Hydrogen-Ion Concentration , India , Kinetics , Lactose/metabolism , Nitrogen/metabolism , Rhodospirillaceae/enzymology , beta-Galactosidase/isolation & purification
14.
J Pharm Anal ; 3(5): 354-359, 2013 Oct.
Article in English | MEDLINE | ID: mdl-29403839

ABSTRACT

In this study, we have demonstrated the conformational changes to DNA induced by abnormal interactions of copper using circular dichroism, in combination with UV-absorbance and fluorescence spectroscopy. Results confirm that binding of copper to bases of DNA in chromatin is concentration dependent. Binding efficiency of Cu2+ ions to DNA is increased in proportion to the degree of unwinding of the double helix induced by denaturation. Altered B-DNA conformation will alter the integrity of DNA which may affect the normal process of DNA replication and transcription. Copper induced DNA damage in the brain may cause neurotoxicity and the neuronal cell death and is implicated in Alzheimer's disease and other neurological disorders.

15.
Biomed Res Int ; 2013: 750187, 2013.
Article in English | MEDLINE | ID: mdl-24455722

ABSTRACT

The present study deals with the production of cold active polygalacturonase (PGase) by submerged fermentation using Thalassospira frigidphilosprofundus, a novel species isolated from deep waters of Bay of Bengal. Nonlinear models were applied to optimize the medium components for enhanced production of PGase. Taguchi orthogonal array design was adopted to evaluate the factors influencing the yield of PGase, followed by the central composite design (CCD) of response surface methodology (RSM) to identify the optimum concentrations of the key factors responsible for PGase production. Data obtained from the above mentioned statistical experimental design was used for final optimization study by linking the artificial neural network and genetic algorithm (ANN-GA). Using ANN-GA hybrid model, the maximum PGase activity (32.54 U/mL) was achieved at the optimized concentrations of medium components. In a comparison between the optimal output of RSM and ANN-GA hybrid, the latter favored the production of PGase. In addition, the study also focused on the determination of factors responsible for pectin hydrolysis by crude pectinase extracted from T. frigidphilosprofundus through the central composite design. Results indicated 80% degradation of pectin in banana fiber at 20 °C in 120 min, suggesting the scope of cold active PGase usage in the treatment of raw banana fibers.


Subject(s)
Fermentation , Pectins/biosynthesis , Polygalacturonase/biosynthesis , Culture Media , Hydrolysis , Rhodospirillaceae/enzymology , Rhodospirillaceae/growth & development
16.
Int J Data Min Bioinform ; 6(4): 396-405, 2012.
Article in English | MEDLINE | ID: mdl-23155770

ABSTRACT

Archaea are ubiquitous in their presence and abundant not only in extreme environments, but also in soil, oceans and freshwater, where they may fulfil a key role in the biogeochemical cycles of the earth. The identification of archaeal genomic signatures elucidates us a measure of distinctiveness of Archaea as a coherent group, although these signatures can differ according to the degree of stringency. The 16S rRNA and the Rad A genes are highly conserved in living organisms and are very useful for the phylogenetic analysis. Phylogenetic trees are constructed using the molecular evolutionary genetics analysis (MEGA) tool by neighbour joining (NJ) method and repeated bootstrapping for 5000 times was performed. The two trees were then compared using the Compare2trees and statistically analysed using the MEGA tool. The two phylogenetic trees show a similarity of 54.9%. In both the trees, the taxon Thaumarchaeota shows a high level of variance. The species Cenarchaeum symbiosum A shows a high level of similarity with the sequences of higher organisms (Euryarcheota-eukaryota), which shows that it has branched away to higher organisms from a closely related Protozoa, Eubacteria ancestor.


Subject(s)
Archaea/genetics , Evolution, Molecular , Genomics/methods , Phylogeny , RNA, Archaeal/genetics , RNA, Ribosomal, 16S/genetics , Genome
17.
Int J Comput Biol Drug Des ; 5(2): 164-79, 2012.
Article in English | MEDLINE | ID: mdl-22854124

ABSTRACT

With the heightened interest in Bacillus anthracis as a potential biological threat agent, novel drug targets identification is of great importance in drug discovery. This study considered a genome-wide approach to identify 270 non-redundant, non-human homologous genes and 103 essential genes of the bacteria as putative drug targets. Sub-cellular localisation of each drug target was annotated using PSORTb 3.0 and confirmation by a hybrid support vector machine analysis identified 16 membrane-bound genes with reliability index ≥4. SPAAN analysis predicted 3 adhesion-like proteins and BLAST against the MEROPS database identified 7 peptidases with inhibitors. As a case study, a homology model was built for the ptsG gene using Modeller 9v8. The work reported here identified a small subset of potential drug targets involved in vital aspects of the metabolism of pathogen, persistence, virulence and cell wall biosynthesis. Thus, this manifold workflow can speed up the process of drug target discovery.


Subject(s)
Anti-Bacterial Agents/pharmacology , Bacillus anthracis/drug effects , Genome, Bacterial , Bacillus anthracis/genetics , Genes, Bacterial
18.
Interdiscip Sci ; 4(2): 142-52, 2012 Jun.
Article in English | MEDLINE | ID: mdl-22843237

ABSTRACT

Tuberculosis (TB), the second most deadly disease in the world is caused by Mycobacterium tuberculosis (Mtb). In the present work a unique enzyme of Mtb orotidine 5' monophosphate decarboxylase (Mtb-OMP Decase) is selected as drug target due to its indispensible role in biosynthesis of pyrimidines. The present work is focused on understanding the structural and functional aspects of Mtb-OMP Decase at molecular level. Due to absence of crystal structure, the 3D structure of Mtb-OMP Decase was predicted by MODELLER9V7 using a known structural template 3L52. Energy minimization and refinement of the developed 3D model was carried out with Gromacs 3.2.1 and the optimized homology model was validated by PROCHECK,WHAT-IF and PROSA2003. Further, the surface active site amino acids were quantified by WHAT-IF pocket. The exact binding interactions of the ligands, 6-idiouridine 5' monophosphate and its designed analogues with the receptor Mtb-OMP Decase were predicted by docking analysis with AUTODOCK 4.0. This would be helpful in understanding the blockade mechanism of OMP Decase and provide a candidate lead for the discovery of Mtb-OMP Decase inhibitors, which may bring insights into outcome new therapy to treat drug resistant Mtb.


Subject(s)
Molecular Docking Simulation , Molecular Dynamics Simulation , Mycobacterium tuberculosis/enzymology , Orotidine-5'-Phosphate Decarboxylase/chemistry , Amino Acid Sequence , Catalytic Domain , Ligands , Molecular Sequence Data , Protein Binding , Protein Structure, Secondary , Reproducibility of Results , Sequence Alignment , Structural Homology, Protein , Thermodynamics
19.
World J Microbiol Biotechnol ; 28(9): 2859-69, 2012 Sep.
Article in English | MEDLINE | ID: mdl-22806727

ABSTRACT

The cold active ß-galactosidase from psychrophilic bacteria accelerate the possibility of outperforming the current commercial ß-galactosidase production from mesophilic sources. The present study is carried out to screen and isolate a cold active ß-galactosidase producing bacterium from profound marine waters of Bay-of-Bengal and to optimize the factors for lactose hydrolysis in milk. Isolated bacterium 3SC-21 was characterized as marine psychrotolerant, halophile, gram negative, rod shaped strain producing an intracellular cold active ß-galactosidase enzyme. Further, based upon the 16S rRNA gene sequence, bacterium 3SC-21 was identified as Thalassospira sp. The isolated strain Thalassospira sp. 3SC-21 had shown the enzyme activity between 4 and 20 °C at pH of 6.5 and the enzyme was completely inactivated at 45 °C. The statistical method, central composite rotatable design of response surface methodology was employed to optimize the hydrolysis of lactose and to reveal the interactions between various factors behind this hydrolysis. It was found that maximum of 80.18 % of lactose in 8 ml of raw milk was hydrolysed at pH of 6.5 at 20 °C in comparison to 40 % of lactose hydrolysis at 40 °C, suggesting that the cold active ß-galactosidase from Thalassospira sp. 3SC-21 would be best suited for manufacturing the lactose free dairy products at low temperature.


Subject(s)
Lactose/metabolism , Milk/chemistry , Rhodospirillaceae/enzymology , Rhodospirillaceae/isolation & purification , beta-Galactosidase/genetics , Animals , Bays/microbiology , Cold Temperature , Hydrogen-Ion Concentration , Hydrolysis , India , RNA, Ribosomal, 16S/genetics , Rhodospirillaceae/classification , Rhodospirillaceae/genetics , Seawater/microbiology , Sodium Chloride/metabolism , beta-Galactosidase/metabolism
20.
J Microbiol Biotechnol ; 22(2): 207-14, 2012 Feb.
Article in English | MEDLINE | ID: mdl-22370350

ABSTRACT

Laccase- and peroxidase-free tyrosinase has commercial importance in the production of L-3, 4-dihydroxyphenylalanine (L-DOPA), which is mainly used in the treatment of Parkinson's disease. In the present study, isolation of an actinomycetes microbial strain capable of producing only tyrosinase is reported. Among all soil isolates, three individual colonies revealed black color around the colony in the presence of tyrosine. Further screening for laccase and peroxidase activities using syringaldazine denoted that one of the isolates, designated as RSP-T1, is laccase and peroxidase negative and produces only tyrosinase. The microbe was authenticated as Streptomyces antibioticus based on 16S ribotyping. Effective growth of this isolate was noticed with the use of medium (pH 5.5) containing casein acid hydrolysate (10.0 g/l), K(2)HPO(4) (5.0 g/l), MgSO(4) (0.25 g/l), L-tyrosine (1.0 g/l), and agar (15 g/l). The scanning electron micrograph depicted that the microbe is highly branched and filamentous in nature. The enzyme production was positively regulated in the presence of copper sulfate. The impact of different fermentation parameters on tyrosinase production depicted that the maximized enzyme titer values were observed when this isolate was grown at 6.5 pH and at 30 degrees C temperature under agitated conditions (220 rpm). Among all the studied physiological parameters, agitation played a significant role on tyrosinase production. Upon optimization of the parameters, the yield of tyrosinase was improved more than 100% compared with the initial yield.


Subject(s)
Monophenol Monooxygenase/metabolism , Streptomyces antibioticus/enzymology , Streptomyces antibioticus/isolation & purification , Cluster Analysis , Copper Sulfate/metabolism , Culture Media/chemistry , DNA, Bacterial/chemistry , DNA, Bacterial/genetics , DNA, Ribosomal/chemistry , DNA, Ribosomal/genetics , Enzyme Activators/metabolism , Hydrazones/metabolism , Hydrogen-Ion Concentration , Laccase/metabolism , Microscopy, Electron, Scanning , Molecular Sequence Data , Peroxidase/metabolism , Phylogeny , RNA, Ribosomal, 16S/genetics , Ribotyping , Sequence Analysis, DNA , Soil Microbiology , Streptomyces antibioticus/cytology , Streptomyces antibioticus/genetics , Temperature , Tyrosine/metabolism
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