Editorial 2 2 Genetically modified organisms (GMOs) are now used in the production of pharmaceutical drugs, gene therapy, and golden rice (containing provitamin A). Although options of buying GMOs in the developing countries are limited now, their availability is expected to increase in these countries in the near future and along with it debates will definitely pursue. The creation and use of GMOs have increased continuously in the past few decades. Since creation of the first recombinant bacteria (E.coli) was reported in 1973, expressing an exogenic Salmonella gene (1), Genentech (now a member of the Roche Group) was the first company to announce the creation of an E.coli strain producing the human protein insulin in 1978 (2). More recently (2009), transgenic animals (forms of GMOs) were approved by the U.S. Food and Drug Administration for the production of the first human biological drug (an anticoagulant) to reduce the probability of blood clots during surgery or childbirth (3). Genetically modified bacteria are now used to produce clotting factors to treat haemophilia (4) and human growth hormone to treat different forms of dwarfism (5). In 2012, genetically modified male mosquitoes containing a lethal gene have been developed and released to the environment to fight Dengue fever which is responsible for the death of thousands of people worldwide (6). Considering the anxiety of public over the safety of these organisms, international organizations such as WHO and FAO have been issuing guidances on assessing the safety of genetically engineered organisms during the past two decades (7). Other societies including International Society for Biosafety Research –ISBR– have also been established to promote scientifically sound biosafety research worldwide. ISBR is organizing an International Symposium on the Biosafety of Genetically Modified Organisms (ISBGMO) which is to be held in St Louis, Missouri, USA, 16-20 September 2012. This symposium is expected to be attended by scientists, regulators and developers from all over the world. Main topics in this symposium will include: Genetically modified animals, biosafety policy and practice, RNAi applications and considerations, genetically modified biofuels, current regulatory challenges, defining environmental harm, concepts and applications for environmental risk assessment and regulatory decision-making, biotechnology and crop improvement in developing countries, and new applications of modern biotechnology in agriculture and future implications. It is very appropriate for the leaders of biotech industry and government regulatory agencies in Iran to attend such conferences for learning, exchanging ideas and participation in the discussions on the safety of GMOs. It looks as certain that the products from GMOs will enter the Iranian animal and food market(s) in the coming years. Therefore, the appropriate regulatory agencies in the Ministries of Health and Agriculture should prepare themselves to scientifically explain to the public the importance of GMOs and why they are safe to use either as food or pharmaceutical drugs. 113 113 Ali M. Ardekani Reproductive Biotechnology Research Center, Avicenna Research Institute, ACECR Reproductive Biotechnology Research Center, Avicenna Research Institute, ACECR Iran Editorial 167.pdf Cohen SN, Chang ACY, Boyer HW, Helling RB. Construction of biologically functional bacterial plasmids in vitro. Proc Natl Acad Sci USA 1973;70(11):3240-3244.##First Successful Laboratory Production of Human Insulin Announced. http://www.gene.com (Accessed July 1, 2012).##FDA Approves Drug From Transgenic Goat Milk. http://www.webcitation.org/ (Accessed July 2012).##Pipe SW. Recombinant clotting factors. Thromb Haemost 2008;99(5):840-850.##Bryant J, Baxter L, Cave CB, Milne R. Recombinant growth hormone for idiopathic short stature in children and ado-lescents. Cochrane Database Syst Rev 2007;(3):CD004440.##Harris AF, Nimmo D, McKemey AR, Kelly N, Scaife S, Donnelly CA, et al. Field performance of engineered male mos-quitoes. Nat Biotechnol 2011;29(11):1034-1037.##Meeting on the safety of foods derived through biotechnology. http://www.who.int/en/ (Accessed July 2012).##

Regulations and Ethical Considerations in Animal Experiments: International Laws and Islamic Perspectives 2 2 Growing usage of animals in the research projects has drawn more attention to their welfare and ethics surrounding this practice. Dissemination of information about the existing ethical consideration and alternatives in animal experiments has two important functions; first, it increases the researcher's awareness of the possible methods of using animals in the experiment, and second, to ensure that potential users are aware of the established alternatives. For example, legislations enacted in many countries during the 1980s state that laboratory animal applications should be reduced, refined and replaced wherever possible according to principles of the 3Rs. Thus, scientists around the world tried to apply the 3Rs in their biomedical researches regarding welfare of the laboratory animals. However, the Qur’an, the holy book of Muslims, and also Hadiths contain the obligatory ways to keep and treat animals since their revelations. According to Islamic viewpoint, animals represent Allah's ability and wisdom, and humans must pay attention to their health and living conditions. Several Islamic manuscripts state that animals have their own position in the creation hierarchy and humans are responsible for supplying minimal facilities and their welfare. This paper has tried to review ethical consideration in animal experiments and regarding Islamic resources in this case to encourage providing comprehensive ethical regulations in animal experiments which its establishment could be beneficial for animal ethics committees or research institutes. 114 120 Mohammad Mehdi Naderi Reproductive Biotechnology Research Center, Avicenna Research institute, ACECR Reproductive Biotechnology Research Center, Avicenna Research institute, ACECR Iran Ali Sarvari Reproductive Biotechnology Research Center, Avicenna Research Institute, ACECR Reproductive Biotechnology Research Center, Avicenna Research Institute, ACECR Iran Alireza Milanifar Nanobiotechnology Research Center, Avicenna Research Institute, ACECR Nanobiotechnology Research Center, Avicenna Research Institute, ACECR Iran Sara Borjian Boroujeni Reproductive Biotechnology Research Center, Avicenna Research Institute, ACECR Reproductive Biotechnology Research Center, Avicenna Research Institute, ACECR Iran Mohammad Mehdi Akhondi Reproductive Biotechnology Research Center, Avicenna Research Institute, ACECR Reproductive Biotechnology Research Center, Avicenna Research Institute, ACECR Iran AnimalsEthicsResearchWelfare 89.pdf Hau J, Van Hoosier GL. Animal research ethics. In: Olsson IAS, Robinson P, Pritchett K, Sandoe P (eds). Handbook of Laboratory Animal Science. Boca Raton: CRC Press; 2003, 13-31.##Hau J, Van Hoosier GL. An overview of global legislation, regulations and policies on the use of animals for scientific research, testing, or education. In: Bayne K, deGreeve P (eds). Handbook of Laboratory Animal Science. USA: Boca Raton; 2003, 31-42.##Suckow MA, Douglas FA, Weichbrod RH. Bioethics and laboratory animal research. In: Gnadt JW, Leland SE (eds). Management of Laboratory Animal Care and Use Programs. Boca Raton: CRC Press; 2002, 92-106.##Hubrecht R, Kirkwood J. Implementing the three Rs in research using animals. In: Richmond J (ed). UFAW Handbook on the Care and Management of Laboratory Animals. Vol. 2. London: Blackwell publishing; 2010, 23-37. ##Naiini A, Rabani M. Institutional Policies and Responsibilities. In: Animal rights according to QURAN viewpoints. Quarterly periodical of Shahed University 1999;26:43-50. Persian.##Barthold SW, bayne AK, Davis MA. Animal care and use program. In: Ballinger MB (ed). Guide for the Care and Use of Laboratory Animals: National Research Council. Washington: National Academy Press; 2011, 11-20. ##Olfert ED, McWilliam AA. Control of Animal Pain in Research, Teaching and Testing. In: Amyx, HL (ed). Guide to the Care and Use of Experimental Animals. Vol. 1. Ottawa: Canadian Council on Animal Care; 1993, 188-189. ##Ruys T. Laboratory Animal Facilities. In: Ruys T (ed). Handbook of Facilities Planning. New York: Van Nostrand Reinhold; 1991, 193-197. ##Fox JG, Anderson LC, Loew FM, Quimby FW. Laws, regulation and policies affecting the use of laboratory animals. In: Anderson LC (ed). La-boratory Animal Medicine. Maryland: Academic Press; 2002, 19-32.##Applied Research Ethics National Association (ARENA) and Office of Laboratory Animal Welfare (OLAW). Institutional Animal Care and Use Committee Guidebook. Bethesda: National government publication; 2002.##National health and medical research council. Australian code of practice for the care and use of animals for scientific purposes. Canberra: Australian Government; 2004, 21-32.##Reilly JS. Animal welfare considerations. In: Reilly JS, Rose MA (eds). Euthanasia of animals used for scientific purposes. Adelaide: Anzccart; 2001, 6-15. ##International Air Transport Association. Live Animals Regulations. 36th ed. Montreal: International Air Transport Association; 2009. ##Flecknell PA. The relief of pain in laboratory ani-mals. Lab Animals 1984;(18):147-160.##Beaver BV, Reed WSL, McKiernan B. Report of the AVMA panel on euthanasia. J Am Vet Med Assoc 2001;218(5):669-696. ##Barthold SW, Bayne KA, Davis MA. Guide for the care and use of laboratory animals. Washington: National Academy Press; 2011. ##Hawkins P. Recognizing and assessing pain, suffering and distress in laboratory animals-a survey of current practice in the UK with recommendations. Lab Anim 2002;36:378-395. ##Mobasher M, Aramesh K, Aldavoud SJ, Ashrafganjooei N, Divsalar K, Phillips CJC, et al. Proposing a national ethical framework for animal research in Iran. Iranian J Publ Health 2008;37 (1):39-46. ##Alhor Alameli M, Vasael Alshieh. Tehran Maktab aleslamieh 8:351-353,397. Arabic.##Ale-Davoud SJ, Javadzade-bolouri A, Sadeghi H, Sabzghabie MAM, Latifi SM, Norouzian R, et al. Preparation of ethic codes for studies on laboratory animals. J Babol Univ Med Sci 2006;8(3):55-64.##Bennison R. Ecological inclusion and nonhuman animals in the Islamic tradition. Koninklijke Brill NV: Leiden; 2003.##

Thioredoxin System: A Model for Determining Novel Lead Molecules for Breast Cancer Chemotherapy 2 2 Background: Thioredoxin reductase 1 (TXNRD1) and thioredoxin interacting protein (TXNIP) also known as thioredoxin binding protein 2 or vitamin D3-upregulated protein 1 are key players in oxidative stress control. Thioredoxin (TRX) is one of the major components of the thiol reducing system and plays multiple roles in cellular processes. Computational analyses of TXNRD1, TXNIP and TRX expressions have not been analyzed in relation to prognosis of breast cancer. High expression of TXNRD1 and low expression of TXNIP are associated with worst prognosis in breast cancer. Methods: Using bioinformatics applications we studied sequence analysis, molecular modeling, template and fold recognition, docking and scoring of thioredoxin as a target. Results: The resultant model obtained was validated based on the templates from I-TASSER server and binding site residues were predicted. The predicted model was used for Threading and Fold recognition and was optimized using GROMACS. The generated model was validated using programs such as Procheck, Ramachandran plot, verify-3d and Errat value from Saves server, and the results show that the model is reliable. Next we obtained small molecules from pubchem and chembank which are databases for selecting suitable ligands for our modeled target. These molecules were screened for docking, using GOLD and scoring was obtained using Chemscore as a scoring function. Conclusion: This study predicted the ligand interaction of four molecules with the minimized protein modeled structure and the best ligand with top scores from about 500 molecules screened. These were 3-hydroxy-2,3-diphenylbutanoic acid, 4-amino-3-pentadecylphenol, 3-(hydroxyimino)-2,4-diphenylbutanenitrile and 2-ethyl-1,2-diphenylbutyl carbamate, which are proposed as possible hit molecules for the drug discovery and development process. 121 130 Kaiser Jamil School of Life Sciences, Centre for Biotechnology and Bioinformatics (CBB), Jawaharlal Nehru Institute of Advanced Studies India Sabeena Muhammed Mustafa Centre for Biotechnology and Bioinformatics (CBB), Jawaharlal Nehru Institute of Advanced Studies India Breast cancerChemotherapySequence analysisThioredoxins 90.pdf Nishiyama A, Matsui M, Iwata S, Hirota K, Masutani H, Nakamura H, et al. Identification of thioredoxin-binding protein-2/vitamin D(3) up-regulated protein 1 as a negative regulator of thioredoxin function and expression. J Biol Chem 1999;274(31):21645-21650. ##Berndt C, Lillig CH, Holmgren A. Thiol-based mechanisms of the thioredoxin and glutaredoxin systems: implications for diseases in the cardiovascular system. Am J Physiol Heart Circ Physiol 2007;292(3):H1227-H1236.##Marks PA. Thioredoxin in cancer—role of histone deacetylase inhibitors. Semin Cancer Biol 2006; 16(6):436-443.##Nalini R, Wilma Delphine Silvia CR, Makhija PM, Uthappa S. Usefulness of serum ca 15.3 and histopathological prognostic indices in breast cancer. Indian J Clin Biochem 2005;20(1):165-168.##Masutani H, Ueda S, Yodoi J. The thioredoxin system in retroviral infection and apoptosis. Cell Death Differ 2005;12(Suppl 1):991-998.##Karlenius TC, Tonissen KF. Thioredoxin and Cancer: A role for Thioredoxin in all states of tumor oxygenation. Cancers 2010;2(2):209-232.##Munemasa Y, Ahn JH, Kwong JM, Caprioli J, Piri N. Redox proteins thioredoxin 1 and thioredoxin 2 support retinal ganglion cell survival in experimental glaucoma. Gene Ther 2009;16(1):17-25.##Zhang Y. I-TASSER server for protein 3D structure prediction. BMC Bioinformatics 2008;9:40.##Kotra S, Madala KK, Jamil K. Homology models of the mutated EGFR and their response towards quinazolin analogues. J Mol Graph Model 2008;27(3):244-254. ##Hess B, Kutzner C, van der Spoel D, Lindahl E. GROMACS 4: algorithms for highly efficient, load-balanced, and scalable molecular simulation. J Chem Theory Comput 2008;4(3):435-447.##Shoichet BK. Virtual screening of chemical libraries. Nature 2004;432:862-865.##Kitchen DB, Decornez H, Furr JR, Bajorath J. Docking and scoring in virtual screening for drug discovery: methods and applications. Nat Rev Drug Discov 2004;3(11):935-949.##Wang Y, Xiao J, Suzek TO, Zhang J, Wang J, Bryant SH. PubChem: a public information system for analyzing bioactivities of small molecules. Nucleic Acids Res 2009;37(Web Server issue):W623-33.##Sudha KN, Shakira M, Prasanthi P, Sarika N, Kumar ChN, Babu PA. Virtual screening for novel COX-2 inhibitors using the ZINC database. Bioinformation 2008;2(8):325-329.##Verdonk ML, Chessari G, Cole JC, Hartshorn MJ, Murray CW, Nissink JW, et al. Modeling water molecules in protein-ligand docking using GOLD. J Med Chem 2005;48(20):6504-6515.##Verdonk ML, Cole JC, Hartshorn MJ, Murray CW, Taylor RD. Improved protein–ligand docking using GOLD. Proteins 2003;52(4):609-623.##Eswar N, Webb B, Marti-Renom MA, Madhusudhan MS, Eramian D, Shen MY, et al. Comparative protein structure modeling using MODELLER. Curr Protoc Protein Sci 2007;Chapter 2:Unit 2.9.##

3’-RACE Amplification of Aminopeptidase N gene From Anopheles stephensi Applicable in Transmission Blocking Vaccines 2 2 Background: Because of the lack of an effective and economical control strategy against malaria (the most devastating infectious disease in developing countries) Transmission-Blocking Vaccines (TBVs) concept has been raised in recent years, promising a more efficient way to malaria control. TBVs aim at interfering and/or blocking pathogen development within the vector, halting transmission to non-infected vertebrate host. Aminopeptidase N (APN) is one of the most potent proteins in parasite development in Anopheles malaria vectors, which is strongly co-localized with human malaria parasites in the mosquito midgut epithelium. Therefore, Aminopeptidase N is one of the best choices for a new TBV. Methods: In this study for the first time we used 3'-RACE to amplify APN gene in Anopheles stephensi (An.stephensi), a major malaria vector in Iran, Indian subcontinent up to China by using different sets of primers including exon junction, conserved and specific region primers. Results: Full length of APN was sequenced stepwise, which could be applied in designing a new regional TBV and act as an essential component of malaria elimination program in An. stephensi distribution areas. Conclusion: Primers design and method modification should be set up exactly in approach based amplifications. From results we came to this conclusion that that 3'-RACE could be applied to amplified key regions which are be-yond reach. 131 141 Hanieh Bokharaei Biology Department, Faculty of Basic Sciences, Science and Research Branch, Islamic Azad UniversityMalaria and Vector Research Group (MVRG), Biotechnology Research Center (BRC), Pasteur Institute of Iran Biology Department, Faculty of Basic Sciences, Science and Research Branch, Islamic Azad University IranIran Abbasali Raz Malaria and Vector Research Group (MVRG), Biotechnology Research Center (BRC), Pasteur Institute of Iran Iran Sedigheh Zakeri Malaria and Vector Research Group (MVRG), Biotechnology Research Center (BRC), Pasteur Institute of Iran Iran Navid Dinparast Djadid Malaria and Vector Research Group (MVRG), Biotechnology Research Center (BRC), Pasteur Institute of Iran Iran 3’-RACEAnopheles stephensiAPNAminopeptidase NMalariaTransmission blocking vaccine 91.pdf Frischknecht F, Martin B, Thiery I, Bourgouin C, Menard R. Using green fluorescent malaria parasites to screen for permissive vector mosquitoes. Malaria J 2006:5;23-29.##Snow RW, Guerra CA, Noor AM, Myint HY, Hay SI. The global distribution of clinical episodes of Plasmodium falciparum malaria. Nature 2005;434(7030):214-217.##Greenwood BM, Bojan K, Whitty CJ, Targett GA. Malaria. Lancet 2005;365(9469):1487-1498.##Bosman A, Mendis KN. A major transition in malaria treatment: the adoption and deployment of artemisinin-based combination therapies. Am J Trop Med Hyg 2007;77(6 Suppl):193-197.##Collins WE, Jeffery GM. Plasmodium malariae: parasite and disease. Clin Microbiol Rev 2007;20(4):579-592.##Rodrigues J, Agrawal N, Sharma A, Malhotra P, Adak T, Chauhan VS, et al. Transcriptional analysis of an immune-responsive serine protease from Indian malarial vector, Anopheles culicifacies. BMC Mol Biol 2007;8:33.##Xu WY, Huang FS, Hao HX, Duan JH, Qiu ZW. Two serine protease from Anopheles dirus heamocytes exhibit changes in transcript abundance after infection of an incompatible rodent malaria parasite, Plasmodium yoelii. Vet Parasit 2006;139(1-3):93-101.##Barillas-Mury C, Kumar S. Plasmodium–mosquito interactions: a tale of dangerous liaisons. Cell Microbiol 2005;7(11):1539-1545.##Ghosh A, Edwards MJ, Jacobs-Lorena M. The journey of the malaria parasite in the mosquito: hopes for the new century. Parasitol Today 2000;16(5):196-201.##Han YS, Thompson J, Kafatos FC, Barillas-Mury C. Molecular interactions between Anopheles stephensi midgut cells and Plasmodium berghei: the time bomb theory of ookinete invasion of mosquitoes. EMBO J 2000;19(22):6030-6040.##Tsuboi T, Cao YM, Hitsumoto Y, Yanagi T, Kanbara H, Torii M. Two antigens on zygotes and ookinetes of Plasmodium yoelii and Plasmodium berghei that are distinct targets of transmission-blocking immunity. Infect Immun 1997;65(6):2260-2264.##Malkin EM, Durbin AP, Diemert DJ, Sattabongkot J, Wu Y, Miura K, et al. Phase 1 vaccine trial of Pvs25H: a transmission blocking vaccine for Plasmodium vivax malaria. Vaccine 2005;23(24):3131-3138.##Kaslow DC, Shiloach J. Production, puriﬁcation and immunogenicity of a malaria transmission-blocking vaccine candidate: TBV25H expressed in yeast and puriﬁed using nickel-NTA agarose. Biotechnology 1994;12(5):494-499.##Kumar N, Ploton I, Koski G, Ann-Lobo C, Contreras C. Malaria transmission-blocking immunity. Identiﬁcation of epitopes and evaluation of immunogenicity. Adv Exp Med Biol 1995;383:65-72.##Saul A. Minimal efficacy requirements for malarial vaccines to significantly lower transmission in epidemic or seasonal malaria. Acta Trop 1993;52(4):283-296.##WHO. Malaria transmission-blocking vaccines: an ideal public good. WHO document no. (TDRyRBMyMALyVACy2000.1) available from WHO, Geneva, and on website available from http:yywww.who.intytdr 2000.##17. Quakyi IA, Carter R, Rener J, Kumar N, Good MF, Miller LH. The 230-kDa gamete surface protein of Plasmodium falciparum is also a target for transmission-blocking antibodies. J Immunol 1987;139 (12):4213-4217.##Duffy PE, Kaslow DC. A novel malaria protein, Pfs28 and Pfs25 are genetically linked and synergistic as falciparum malaria transmission-blocking vaccines. Infect Immun 1997;65(3):1109-1113.##Hisaeda H, Stowers AW, Tsuboi T, Collins WE, Sattabongkot JS, Suwanabun N, et al. Antibodies to malaria vaccine candidates Pvs25 and Pvs28 completely block the ability of Plasmodium vivax to infect mosquitoes. Infect Immun 2000;68 (12):6618-6623.##Sattabongkot J, Tsuboi T, Hisaeda H, Tachibana M, Suwanabun N, Rungruang T, et al. Blocking of transmission to mosquitoes by antibody to Plasmodium vivax malaria vaccine candidates Pvs25 and Pvs28 despite antigenic polymorphism in field isolates. Am J Trop Med Hyg 2003;69 (5):536-541.##Rosenberg R, Koontz LC, Alston K, Friedman FK. Plasmodium gallinaceun: erythrocyte factor essential for zygote infection of Aedes aegypti. Exp Parasitol 1984;57 (2):158-164.##Dinglasan RR, Fields I, Shahabuddin M, Azad AF, Sacci JB Jr. Monoclonal antibody MG96 completely blocks Plasmodium yoelii development in Anopheles stephensi. Infect Immun 2003;71(12):6995-7001.##Dinglasan RR, Valenzuela JG, Azad AF. Sugar epitopes as potential universal disease transmission blocking targets. Insect Biochem Mol Biol 2005;35 (1):1-10.##Dinglasan RR, Alaganan A, Ghosh A, Saito A, van Kuppevelt T, Jacobs-Lorena M. Plasmodium falciparum ookinetes require mosquito midgut chondroitin sulfate proteoglycans for cell invasion. Proc Natl Acad Sci USA 2007;104(40):15882-15887.##Rosenfeld A, Vanderberg JP. Plasmodium berghei: induction of aminopeptidase in malaria-resistant strain of Anopheles gambiae. Exp Parasitol 1999;93(2):101-104.##Taylor A. Aminopeptidases: structure and function. FASEB J 1993;7(2):290-298.##Hooper NM. Families of zinc metalloproteases. FEBS Lett 1994;354(1):1-6.##Mohammadzadeh Hajipirloo H, Edrissian GhH, Nateghpour M, Basseri H, Eslami MB. Effects of anti-mosquito salivary glands and deglycosylated midgut antibodies of Anopheles stephensi on fecundity and longevity. Iranian J Publ Heal 2005;34(4):8-14.##Dinglasan RR, Kalume DE, Kanzok SM, Ghosh AK, Muratova O, Pandey A, et al. Disruption of Plasmodium falciparum development by antibodies against a conserved mosquito Midgut antigen. Proc Natl Acad Sci U S A 2007;104(33):13461-13466.##Klemba M, Gluzman I, Goldberg DE. A Plasmodium falciparum dipeptidyl Aminopeptidase I participates in vacuolar hemoglobin degradation. J Biol Chem 2004;279(41):43000-43007.##Taylor A, Carpenter FH, Wlodawer A. Leucine aminopeptidase N (bovine lens): an electron microscopic study. J Ultrastruct Res 1979;68 (1):92-100.##Nishizawa R, Saino T, Takita T, Suda H, Aoyagi T. Synthesis and structure-activity relationships of bestatin analogues, inhibitors of aminopeptidase. J Med Chem 1977;20(4):510-515.##Allen MP, Yamada AH, Carpenter FH. Kinetic parameters of metal-substituted leucine aminopeptidase from bovine lens. Biochemistry 1983;22(16):3778-3783.##Garcia-Alvarez N, Cueva R, Suarez-Rendueles P. Molecular cloning of soluble aminopeptidases from Saccharomyces cerevisiae. Sequence analysis of aminopeptidase ysc II, a putativezinc-metallo peptidase. Eur J Biochem 1991;202(3):993-1002.##Xu W, Li Q. Progress in the development of aminopeptidase N (APN/CD13) inhibitors. Curr Med Chem Anticancer Agents 2005;5(3):435-476.##Ito K, Nakajima Y, Onohara Y, Takeo M, Nakashima K, Matsubara F, et al. Crystal structure of aminopeptidase N (proteobacteria alanyl aminopeptidase) from Escherichia coli and conformational change of methionine 260 involved in substrate recognition. J Biol Chem 2006;281(44):33664-33676.##

Development of an Immunoaffinity Method for Purification of Streptokinase 2 2 Background: Streptokinase is a potent activator of plasminogen to plasmin, the enzyme that can solubilize the fibrin network in blood clots. Streptokinase is currently used in clinical medicine as a thrombolytic agent. It is naturally secreted by β-hemolytic streptococci. Methods: To reach an efficient method of purification, an immunoaffinity chromatography method was developed that could purify the streptokinase in a single step with high yield. At the first stage, a CNBr-Ac-tivated sepharose 4B-Lysine column was made to purify the human blood plasminogen. The purified plasminogen was utilized to construct a column that could purify the streptokinase. The rabbit was immunized with the purified streptokinase and the anti-streptokinase (IgG) purified on another streptokinase substituted sepharose-4B column. The immunoaffinity column was developed by coupling the purified anti-Streptokinase (IgG) to sepharose 6MB–Protein A. The Escherichia coli (E.coli) BL21 (DE3) pLysS strain was transformed by the recombinant construct (cloned streptokinase gene in pGEX-4T-2 vector) and gene expression was induced by IPTG. The expressed protein was purified by immunoaffinity chromatography in a single step. Results: The immunoaffinity column could purify the recombinant fusion GST-SK to homogeneity. The purity of streptokinase was confirmed by SDS-PAGE as a single band of about 71 kD and its biological activity determined in a specific streptokinase assay. The yield of the purification was about 94%. Conclusion: This method of streptokinase purification is superior to the previous conventional methods. 142 147 Zohreh Karimi Department of Biology, Faculty of Science, Alzahra University Department of Biology, Faculty of Science, Alzahra University Iran Mohammad Babashamsi Department of Immunochemistry, Monoclonal Antibody Research Center, Avicenna Research Institute, ACECR Department of Immunochemistry, Monoclonal Antibody Research Center, Avicenna Research Institute, ACECR Iran Ezat Asgarani Department of Biology, Faculty of Science, Alzahra University Department of Biology, Faculty of Science, Alzahra University Iran Ali Salimi Department of Immunochemistry, Monoclonal Antibody Research Center, Avicenna Research Institute, ACECR Department of Immunochemistry, Monoclonal Antibody Research Center, Avicenna Research Institute, ACECR Iran ChromatographyPurificationStreptokinaseThrombolytic agent 92.pdf Banerjee A, Chistic Y, Banerjee UC. Streptokinase, a clinically useful thrombolytic agent. Biotechnol Adv 2004;22(4):287-307. ##Tillet WS, Garner RL. The fibrinolytic activity of hemolytic streptococci. J Exp Med 1933;58(4):485-502.##Castellino FJ. A unique enzyme — protein substrate modifier reaction: plasmin/streptokinase interaction. Trends Biochem Sci 1979;4(1):1-5.##Rodriguez P, Fuentes P, Barro M, Alvarez JG, Muñoz E, Collen D, et al. Structural domains of streptokinase involved in the interaction with plasminogen. Eur J Biochem 1995;229(1):83-90.##Jackson KW, Tang J. Complete amino acid sequence of streptokinase and its homology with serine proteases. Biochemistry 1982;21(26):6620-6625.##Blatt WF, Segal H and Gray JL. Purification of streptokinase and human plasmin and their interaction. Thromb Diath Haemorrh 1964;11:393-403.##Jeong IK, Young EL, Jeung SJ, Myung B. Purification of streptokinase by affinity chromatography using human plasminogen. Korean Biochem J 1993;26(2):172-175.##Karush F, Lacocca VF, Harris TN, Growth of group a hemolytic streptococcus in the steady state. J Bacteriol 1956;72(3):283-294. ##Ogburn CA, Harris TN, Harris S. Extracellular antigens in steady-state cultures of the hemolytic streptococcus: production of proteinase at low pH. J Bacteriol 1958;76(2):142-151.##Babashamsi M, Razavian MH, Nejadmoghaddam MR. Production and purification of streptokinase by protected affinity chromatography. Avicenna J Med Biotech 2009;1(1):47-51.##Steiner K, Malke H. Dual control of streptokinase and streptolysin S production by the covRS and fasCAX two-component regulators in Streptococcus dysgalactiae subsp. equisimilis. Infect Immun 2002;70(7):3627-3636.##Kaur J, Rajamohan G, Dikshit KL. Cloning and characterization of promoter-active DNA sequences from Streptococcus equisimilis. Curr Microb 2007;54(1):48-53.##Nejadmoghaddam MR, Modarresi MH, Babashamsi M, Chamankhah M. Cloning and over expression of active recombinant fusion streptokinase: a new approach to facilitate purification. Pak J Biol Sci 2007;10(13):2146-2151. ##Jackson KW, Esmon N, Tang J. Streptokinase and staphylokinase. Methods Enzymol 1981;80:387-394. ##De Renzo EC, Suteri PK, Hutchings BL, Bell PH. Preparation and certain properties of highly purified streptokinase. J Biol Chem 1967;242(3):533-542.##Taylor FB, Botts J. Purification and characterization of streptokinase with studies of streptokinase activation of plasminogen. Biochemistry 1968;7:232-242.##Einarsson M, Skoog B, Forsberg B, Einarsson R. Characterization of highly purified native streptokinase and altered streptokinase after alkaline treatment. Biochimica et Biophysica Acta 1979;568(1):19-29.##Castellino FJ, De Renzo EC, Suteri PK, Hutchings BL, Bell PH. Preparation and certain properties of highly purified streptokinase. J Biol Chem 1967;242;533-542.##Karimi Z, Babashamsi M, Asgarani E, Niakan M, Salimi A. Fermentation, fractionation and purification of streptokinase by chemical reduction method. Iran J Microbiol 2011;3(1):42-46.##

Reduction of Sodium Arsenite-Mediated Adverse Effects in Mice using Dietary Supplementation of Water Hyacinth (Eichornia crassipes) Root Powder 2 2 Background: In this study, we evaluated the protective effects of water Hyacinth Root Powder (HRP) on arsenic-mediated toxic effects in mice. Methods: Swiss albino mice, used in this study, were divided into four different groups (for each group n=5). The control group was supplied with normal feed and water, Arsenic group (As-group) was supplied with normal feed plus arsenic (sodium arsenite)-containing water, and arsenic+hyacinth group (As+Hy group) was supplied with feed supplemented with HRP plus arsenic water. The remaining Hy-group was supplied with feed supplemented with HRP plus normal water. Results: Oral administration of arsenic reduced the normal growth of the mice as evidenced by weight loss. Interestingly, tip of the tails of these mice developed wound that caused gradual reduction of the tail length. Supplementation of HRP in feed significantly prevented mice growth retardation and tail wounding in As+Hy group mice. However, the growth pattern in Hygroup mice was observed to be almost similar to that of the control group indicating that HRP itself has no toxic or negative effect in mice. Ingested arsenic also distorted the shape of the blood cells and elevated the serum enzymes such as lactate dehydrogenase (LDH), alkaline phosphatase (ALP) and serum glutamic pyruvic transaminase (SGPT). Importantly, elevation of these enzymes and distortion of blood cell shape were partially reduced in mice belong to As+Hy group, indicating HRP-mediated reduction of arsenic toxicity. Conclusion: Therefore, the preventive effect of hyacinth root on arsenic-poisoned mice suggested the future application of hyacinth to reduce arsenic toxicity in animal and human. 148 154 Rim Sabrina Jahan Sarker Department of Genetic Engineering and Biotechnology, University of Dhaka Department of Genetic Engineering and Biotechnology, University of Dhaka Bangladesh Nazmul Ahsan Department of Genetic Engineering and Biotechnology, University of Dhaka Department of Genetic Engineering and Biotechnology, University of Dhaka Bangladesh Khaled Hossain Department of Biochemistry and Molecular Biology, Rajshahi University Department of Biochemistry and Molecular Biology, Rajshahi University Bangladesh Paritosh Kumar Ghosh Department of Pathology, Bangladesh Medical College Bangladesh Chowdhury Rafiqul Ahsan Department of Microbiology, University of Dhaka Department of Microbiology, University of Dhaka Bangladesh Anwarul Azim Akhand Department of Genetic Engineering and Biotechnology, University of Dhaka Department of Genetic Engineering and Biotechnology, University of Dhaka Bangladesh ArsenicBlood cell morphologyGrowth retardationMiceWater hyacinth 93.pdf Khan MMH, Aklimunnessa K, Ahsan N, Kabir M, Mori M. 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The Anticonvulsant and Neuroprotective Effects of Walnuts on the Neurons of Rat Brain Cortex 2 2 Background: Epilepsy is a chief communal health problem. Antiepileptic drugs only provide symptomatic treatment. Walnut Kernels (WK) have high concentrations of phenolic compounds, which have beneficial effects on human health because of their antioxidant and anti-atherogenic properties. The present study was designed to evaluate the efficacy of WK supplementation for the prevention of experimental epilepsy in male rats. Methods: Wistar adult male rats were divided into three groups: a control group (PTZ injection, fed with ordinary food), experimental group (PTZ injection, fed with WK) and a sham group (no PTZ injection, only for histological studies). Pentylenetetrazole (PTZ) was administered after the prescribed time. Results: WKs displayed anti-epileptogenic properties, and WK supplementation was associated with increased seizure threshold and reduced mortality in the experimental group versus controls. Conclusion: Use of WK may be helpful in prevention of PTZ-induced seizure and its further neurodegeneration in male rats. 155 158 Majid Asadi-Shekaari Neuroscience Research Center, Kerman University of Medical Sciences, Neuroscience Research Center, Kerman University of Medical Sciences, Iran Taj Pari Kalantaripour Faculty of Midwifery and Nursing, Islamic Azad University, Kerman branch Faculty of Midwifery and Nursing, Islamic Azad University, Kerman branch Iran Fatemeh Arab Nejad Member of Student Research Committee, Kerman University of Medical Sciences Member of Student Research Committee, Kerman University of Medical Sciences Iran Elaheh Namazian Member of Student Research Committee, Kerman University of Medical Sciences Member of Student Research Committee, Kerman University of Medical Sciences Iran Azam Eslami Department of Physiology, Science and Research Branch, Islamic Azad University Department of Physiology, Science and Research Branch, Islamic Azad University Iran AntioxidantsNeuroprotectiveRatSeizures 94.pdf Browne TR, Holmes GL. 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