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Purification of full-length human Pregnane and Xenobiotic Receptor:polyclonal antibody preparation for immunological characterization

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Cell Research 2005年第10期 39-49页

作者： Mallampati SARADHI,Biji kRISHNA,Gauranga MUkHOPADHYAY,Rakesh k tyagi Special Centre for Molecular Medicine Jawaharlal Nehru University New Delhi 110067 India Special Centre for Molecular Medicine Jawaharlal Nehru University New Delhi 110067 India Special Centre for Molecular Medicine Jawaharlal Nehru University New Delhi 110067 India Special Centre for Molecular Medicine Jawaharlal Nehru University New Delhi 110067 India

Pregnane and Xenobiotic Receptor (PXR; or Steroid and Xenobiotic Receptor, SXR), a new member of the nuclear receptor superfamily, is thought to modulate a network of genes that are involved in xenobiotic metabolism and elimination. To further explore the role of PXR in body’s homeostatic mechanisms, we for the first time, report successful prokary- otic expression and purification of full-length PXR and preparation of polyclonal antibody against the whole protein. The full-length cDNA encoding a 434 amino acids protein was sub-cloned into prokaryotic expression vector, pET-30b and transformed into E. coli BL21(DE3) cells for efficient over expression. The inclusion body fraction, containing the expressed recombinant protein, was purified first by solubilizing in sarcosine extraction buffer and then by affinity column chromatography using Ni-NTA His-Bind matrix. The efficacy of anti-PXR antibody was confirmed by immunocytology, Western blot analysis, EMSA and immunohistochemistry. The antibody obtained was capable of detecting human and mouse PXR with high specificity and sensitivity. Immunofluorescence staining of COS-1 cells transfected with human or mouse PXR showed a clear nuclear localization. Results from immunohistochemistry showed that level of PXR in liver sections is immunologically detectable in the nuclei. Similar to exogenously transfected PXR, Western blot analysis of cell extract from HepG2 and COLO320DM cells revealed a major protein band for endogenous PXR having the expected molecular weight of 50 kDa. Relevance of other immunodetectable bands with reference to PXR isoforms and current testimony are evaluated. Advantages of antibody raised against full-length PXR protein for functional characterization of receptor is discussed and its application for clinical purposes is envisaged.

关键词： Pregnane and Xenobiotic Receptor Steroid and Xenobiotic Receptor prokaryotic expression polyclonal antibody isoforms.

Evaluation of anti-apoptotic activity of different dietary antioxidants in renal cell carcinoma against hydrogen peroxide

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Asian Pacific Journal of Tropical Biomedicine 2011年第1期1卷 57-63页

作者： Garg Neeraj k Mangal Sharad Sahu Tejram Mehta Abhinav Vyas Suresh P tyagi Rajeev k Department of Pharmaceutical Sciences Dr.H.S.Gour University Biologie et genetiqjue du paludisme Institut Pasteur28 Rue Du Docteur Roux75724 Paris Cedex 15France Biomedical Parasitology Unit Pasteur Institute

Objective:To evaluate the anti-apoptotic and radical scavenging activities of dietary phenolics, namely ascorbic acid,a-tocopherol acetate,citric acid,salicylic acid,and estimate H_2O_2induced apoptosis in renal cell carcinoma ***:The intracellular antioxidant potency of antioxidants was investigated.H_2O-2-induced apoptosis in RCC-26 was assayed with the following parameters:cell viability(%apoptosis),nucleosomal damage and DNA fragmentation, bcl-2 levels and flow cytometery analysis(ROS production evaluation).Results:Ine anticancer properties of antioxidants such as ascorbic acid,a- tocopherol acetate,citric acid,salicylic acid with perdurable responses were *** was observed that these antioxidants had protective effect(anti-apoptotic activity) against hydrogen peroxide(H_2O_2) in renal cell carcinoma(RCC-26) cell ***:This study reveals and proves the anticancer ***,in cancer cell lines anti-apoptotic activity can indirectly reflect the cancer promoter activity through radicals scavenging,and significantly protect nucleus and bcl-2.

关键词： Antioxidants Anticancer Apoptosis ROS Hydrogen peroxide RCC-26

Provenance variations in proximate principles,mineral matter,total phenols and phytochemicals of Melia dubia drupes:an unexplored alternate livestock feed stock

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Journal of Forestry Research 2021年第1期32卷 119-131页

作者： M.L.Sukhadiya N.S.Thakur V.R.Patel R.P.Gunaga V.B.kharadi k.k.tyagi Susheel Singh Department of Silviculture and Agroforestry College of ForestryNavsari Agricultural UniversityNavsariGujarat 396450India Department of Animal Nutrition Vanbandhu College of Veterinary Sciences and Animal HusbandryNavsari Agricultural UniversityNavsariGujarat 396450India Food Quality Testing Laboratory N.M.College of AgricultureNavsari Agricultural UniversityNavsariGujarat 396450India Sardar Vallabhbhai Patel University of Agriculture and Technology MeerutIndia

This research was carried out on nine provenances from the north Western Ghats to quantify proximate principles,mineral matter,total phenols and other phytochemicals in Melia dubia drupe *** biometric,biomass attributes,proximate principles,mineral matter and total phenols diff ered signifi cantly among the *** average drupe length and width was 26.5 and 22.3 mm,*** drupe and pulp fresh weight,pulp dry weight and stone weight were 783.1,552.1,165.1,and 179.8 g/100 drupes,***,dry matter and moisture per cent were 70.5%,29.9%and 70.0%,*** crude fi bre,protein,ether extract,total ash and nitrogen-free extract were 8.3%,7.6%,5.1%,6.3%,and 72.6%,*** Ca,Mg,P and k levels were 0.5%,0.2%,0.2%,and 2.0%,*** phenols were 0.4%.Cluster analysis showed that the study area may be divided into three distinct provenances,each having diff erent drupe traits and proximate composition,including mineral ***-chromatography mass-spectrometry analysis revealed 27 phytochemicals,many of which possess benefi cial biological *** implications of this provenance variation study should be useful for future improvement in terms of better drupe pulp nutrition for livestock *** study suggests that *** pulp could be categorized as a good feed source with low fi bre,energy,fat and mineral *** proximate parameters,mineral matter content and total phenols are within prescribed limits.

关键词： Melia dubia Drupe pulp Proximate principles Minerals Phytochemicals

Antioxidant Machinery Related to Decreased MDA Generation by Thymus Algeriensis Essential Oil-induced Liver and kidney Regeneration

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Biomedical and Environmental Sciences 2016年第9期29卷 639-649页

作者： Guesmi Fatma Amit k tyagi Bellamine Houda Landoulsi Ahmed Laboratory of Biochemistry and Molecular Biology Faculty of Sciences of Bizerte University of Carthage 1054 Tunisia Department of Experimental Therapeutics The University of Texas M. D. Anderson Cancer Center 1515 Holcombe Boulevard Houston TX 77030 USA Service of Anatomo-Pathology of Menzel Bourguiba Bizerte 1054 Tunisia

Objective This study was conducted to determine the histopathological and biochemical effects of Thymus algeriensis essential oil （TEO） on hydrogen peroxide （H2O2）-induced oxidative stress in liver and kidney tissues of rats. Methods Rats were treated in six groups and were exposed for 2 weeks to low （LD; 100 μmol/L） and high doses （HD; 1 mmol/L） of H2O2 in the presence or absence of TEO （180 mg/kg）. Liver and kidney atrophy was measured by using biochemical and histopathological assays. Results Our study demonstrated that H2O2 induced liver and kidney atrophy, as evidenced by the significant elevation of serum aminotransferase, urea, and creatinine levels compared with those in the control rats. Urea levels were estimated by evaluating the activity of serum urease that hydrolyzes urea into CO2 and ammonia. However, TEO treatment significantly alleviated oxidative stress in the H2O2-induced liver and kidney toxicity model by reducing the levels of malondialdehyde concomitantly with marked elevations in superoxide dismutase, catalase, glutathione peroxidase, and glutathione S-transferase, as well as decrease in glutathione activity. Conclusion Our data demonstrated that TEO protected against H202 toxicity by decreasing oxidant levels and DNA damage, as well as increasing antioxidant levels, indicating that TEO has a spectrum of antioxidant and DNA-protective properties.

关键词： Liver and kidney atrophy Hydrogen peroxide Thymus algeriensis Antioxidants Toxicity

Golgi localization and dynamics of hyaluronan binding protein 1 (HABP1/p32/C1QBP) during the cell cycle

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Cell Research 2005年第3期15卷 183-186页

作者： Aniruddha SENGUPTA,Bhaswati BANERJEE,Rakesh k. tyagi,kasturi DATTA Biochemistry Laboratory School of Environmental Sciences Jawaharlal Nehru University New Delhi-110067 India Biochemistry Laboratory School of Environmental Sciences Jawaharlal Nehru University New Delhi-110067 India Special Centre for Molecular Medicine Jawaharlal Nehru University New Delhi India Biochemistry Laboratory School of Environmental Sciences Jawaharlal Nehru University New Delhi-110067 India2Special Centre for Molecular Medicine Jawaharlal Nehru University New Delhi India

Hyaluronan binding protein 1 (HABP1) is a negatively charged multifunctional mammalian protein with a unique structural fold. Despite the fact that HABP1 possesses mitochondrial localization signal, it has also been localized to other cellular compartments. Using indirect immunofluorescence, we examined the sub-cellular localization of HABP1 and its dynamics during mitosis. We wanted to determine whether it distributes in any distinctive manner after mitotic nuclear envelope disassembly or is dispersed randomly throughout the cell. Our results reveal the golgi localization of HABP1 and demonstrate its complete dispersion throughout the cell during mitosis. This distinctive distribution pattern of HABP1 during mitosis resembles its ligand hyaluronan, suggesting that in concert with each other the two molecules play critical roles in this dynamic process.

关键词： Hyaluronan Binding Protein 1 (HABP1) hyaluronan sub-cellular localization golgi mitosis.

The Receptor-Like Cytoplasmic kinase （OsRLCk） Gene Family in Rice： Organization, Phylogenetic Relationship, and Expression during Development and Stress

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Molecular Plant 2008年第5期1卷 732-750页

作者： Shubha Vij Jitender Giri Prasant kumar Dansana Sanjay kapoor Akhilesh k. tyagi Interdisciplinary Centre for Plant Genomics and Department of Plant Molecular Biology University of Delhi South Campus Benito Juarez Road New Delhi 110021 India

Receptor-like cytoplasmic kinases （RLCks） in plants belong to the super family of receptor-like kinases （RLks）. These proteins show homology to RLks in kinase domain but lack the transmembrane domain. Some of the functionally characterized RLCks from plants have been shown to play roles in development and stress responses. Previously, 149 and 187 RLCk encoding genes were identified from Arabidopsis and rice, respectively. By using HMM-based domain structure and phylogenetic relationships, we have identified 379 OsRLCks from rice. OsRLCks are distributed on all 12 chromosomes of rice and some members are located on duplicated chromosomal segments. Several OsRLCks probably also undergo alternative splicing, some having evidence only in the form of gene models. To understand their possible functions, expression patterns during landmark stages of vegetative and reproductive development as well as abiotic and biotic stress using microarray and MPSS-based data were analyzed. Real-time PCR-based expression profiling for a selected few genes confirmed the outcome of microarray analysis. Differential expression patterns observed for majority of OsRLCks during development and stress suggest their involvement in diverse functions in rice. Majority of the stress-responsive OsRLCks were also found to be localized within mapped regions of abiotic stress QTLs. Outcome of this study would help in selecting organ/development stage specific OsRLCk genes/targets for functional validation studies.

关键词： abiotic stress biotic stress genome-wide analysis kinase rice RLCk.

Development of Advanced Biomass Cookstove and Performance Comparisons Using the Modified Star Rating Methodology

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Energy Engineering 2021年第5期118卷 1237-1251页

作者： Himanshu kunwar Pal Sanjeev Jain S.k.tyagi Department of Energy Science and Engineering Indian Institute of Technology DelhiHauz KhasNew Delhi110016India Department of Chemical Engineering Indian Institute of Technology DelhiHauz KhasNew Delhi110016India Department of Mechanical Engineering Indian Institute of Technology DelhiHauz KhasNew Delhi110016India

A disruptive approach to a fundamental process has been applied in a biomass combustion device with two variable speed fans to supply air for gasification and another for combustion processes,***,the preheating of secondary air,required for combustion process was also ensured through annulus chamber before being fed into the combustion *** turbulent flow and homogenous mixing were also ensured by controlling the flow rate resulting in the reduced emissions of carbon monoxide(CO)and fine particulate matter(PM 2.5,particulate matter having aerodynamic diameterkstove models,and very close to that of a liquefied petroleum gas(LPG)***,the comparative analysis based on the modified star rating of total 15(14 are biomass and another LPG)cookstove models tested using the same standard methodology has been done and presented in this *** on the star rating,the performance of the LPG stove was found to be best and assigned as a 5-star product followed by the IITD model(4-star),while the other 13 models got different ratings starting from 1-star to 3-star,***,the thermal performance of the IITD cookstove model is found to be the highest,while the emission characteristics are found to be the least among all biomass cookstove models,presented here.

关键词： Star rating biomass pellet cookstove thermal performance fine particulate matter(PM2.5) carbon monoxide

Variation in Parameters of Ambient Air Quality in National Capital Territory (NCT) of Delhi (India)

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Atmospheric and Climate Sciences 2015年第1期5卷 13-22页

作者： Anunay A. Gour Santosh k. Singh Sushil k. tyagi Anubha Mandal Department of Environmental Engineering Delhi Technological University (Formerly Delhi College of Engineering) Delhi India Central Pollution Control Board (CPCB) Delhi India

Air Pollution is an escalating problem worldwide fuelled by increasing anthropogenic activities, speedy development, rapid industrialization, transportation, superfluous use of fossil fuel consumption, increasing global power needs, etc. Also the diverse meteorological factors influence our environment to a great extent. Population explosion has led to remarkable increase in vehicular population. The use solely depends upon the lifestyle of the citizen. Any change in the lifestyle has a remarkable effect on the air we breathe. A comprehensive database for pollution levels from the year 2006 to 2010 was analyzed for a main traffic intersection and a non-traffic area in Delhi (India). The paper asserts that pollutant concentration levels on weekend are lower than those on weekdays (increase on Sundays with respect to weekdays is CO 2.98 times, PM2.5 1.18 times, NO 1.62 times, NO2 5.76 times, SO2 1.27 times and O3 1.15 times). The pollution levels have been found to attenuate on public holidays and weekends. Pollution can hence be related to the weekly cycle of human activities. Pollutions due to vehicular emissions on roads, rails, air or water all serve as major sources of pollution. Hence, viably possible solutions for pollution control can also be achieved by controlling the magnitude of traffic flow on roads.

关键词： Ambient Air Quality Emission Seasonal Variation Weekend Effect Delhi

Prevention of H2O2 Induced Oxidative Damages of Rat Testis by Thymus algeriensis

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Biomedical and Environmental Sciences 2016年第4期29卷 275-285页

作者： Fatma Guesmi Hamida Beghalem Amit k tyagi Manel Ben Ali Ramla Ben Mouhoub Houda Bellamine Ahmed Landoulsi Laboratory of Biochemistry and Molecular Biology Faculty of Sciences of Bizerte Zarzouma Bizerte 7021 Tunisia Cytokine Research Laboratory Department of Experimental Therapeutics The University of Texas M. D. Anderson Cancer Center 1515 Holcombe Boulevard Houston TX 77030 USA Laboratory of Biomonitoring of the Environment (LRO1/ES14) Faculty of Sciences of Bizerte Zarzouma Bizerte 7021 Tunisia Service of Anatomo-Pathology of Menzel Bourguiba Bizerte 7050 Tunisia

Objective We evaluate the effects of Thymus algeriensis （TEO） against hydrogen peroxide （H202） toxicity on body and testis weight, testis sperm count, testis lipid peroxidation, and antioxidant enzyme activities in rats. Methods Rats were treated with low （LD） and high dose （HD） of H202 （0.1 and 1 mmol/L） in the presence or absence of TEO （150 mg/kg）. Results The results exhibited a significant decrease in body weight and testis weight, in total sperm number decrease （P〈0.05）, sperm motility and percentage of sperm viability, leading to complete arrest, in sperm flagellar beat frequency by the gavage of 1 mmol/L H202 compared to controls. The administration of H2O2 resulted in a significant reduction in testis GSH, GPx, CAT, SOD, and GST activity and significant increase （P〈0.05） in MDA concentration compared with the untreated control animals. TEO pre-treatment protected testis from the H2O2 generated oxidative stress. These results were confirmed by histological architecture examinations. Conclusion H2O2 has the ability to alter the sperm function, characteristics and development of testis. However, TEO is an efficient natural agent, which can prevent the testis from H2O2-induced oxidative damage in rats.

关键词： Testis Hydrogen peroxide TEO Antioxidants Infertility