National Institute of Genetic Engineering and Biotechnology of IranIranian Journal of Biotechnology1728-30431220030401Vacuolating Cytotoxin of Helicobacter pylori73816891ENYeganeh TalebkhanBiotechnology Research Center, Pasteur Institute of Iran, Tehran, Iran.Marjan MohammadiBiotechnology Research Center, Pasteur Institute of Iran, Tehran, Iran.Journal Article20030401Vacuolating cytotoxin (VacA) is one of the most important virulence factors of H. pylori (Hp), which is<br />the only toxic protein that is secreted from Hp cell into the culture supernatant. The effects of VacA on<br />eukaryotic systems is the subject of many previous and on going research studies. Intracellular targets<br />for this toxin include: late endosomal and lysosomal compartments, mitochondria, cell-cell junctions and<br />phospholipid bilayers. Its effects on these targets include vacuolation of late endosomal and lysosomal<br />compartments, apoptosis and channel formation, which result in the increase of ion uptake specially<br />anions. The aim of this review is to increase the perception on this toxin and its functions.<br /><br />National Institute of Genetic Engineering and Biotechnology of IranIranian Journal of Biotechnology1728-30431220030401Cloning and Expression of Human Gamma-Interferon cDNA in E. coli87946872ENM ArbabiNational Research Center for Genetic Engineering and Biotechnology (NRCGEB), P. O. Box: 14155-6343,
Tehran, Iran.F AlastiNational Research Center for Genetic Engineering and Biotechnology (NRCGEB), P. O. Box: 14155-6343,
Tehran, Iran.MH SanatiNational Research Center for Genetic Engineering and Biotechnology (NRCGEB), P. O. Box: 14155-6343,
Tehran, Iran.S HosseiniBiotechnology Research Center, P.O. Box: 19395-1949, Tehran, Iran.A DeldarBiotechnology Research Center, P.O. Box: 19395-1949, Tehran, Iran.N MaghsoudiBiotechnology Research Center, P.O. Box: 19395-1949, Tehran, Iran.Journal Article20030401Prior to the production of human gamma interferon using recombinant DNA technology, it had been produced<br />mainly upon mitogenic induction of lymphocytes in very low amounts, which evidently hampered<br />its characterization and its medical applications. The recombinant gamma interferons produced in larger<br />quantities in prokaryotic systems retain their biological activities, and can be used clinically in the treatment<br />of various viral, neoplastic and immunosuppressed conditions or diseases. In this study, a cDNA<br />sequence coding for human gamma interferon was synthesized from mRNA template extracted from<br />induced human T lymphocytes. The cDNA was then amplified by PCR, cloned in an expression vector,<br />and transformed into Escherichia coli. The polypeptide produced through the expression of this DNA<br />sequence in E. coli showed immunological and chemical properties resembling authentic human IFN-γ.<br /><br />National Institute of Genetic Engineering and Biotechnology of IranIranian Journal of Biotechnology1728-30431220030401Isolation and Genetic Fingerprinting of Pseudomonas aeruginosa from Iranian Patients with Cystic Fibrosis Using RAPD-PCR951006873ENFereshteh EftekharBiology Department, Faculty of Science, Shahid Beheshti University. Evin, Tehran, Iran.Farkhondeh RostamizadehBiology Department, Faculty of Science, Shahid Beheshti University. Evin, Tehran, Iran.Ahmad KhodadadChildren’s Hospital Medical Center, Tehran University of Medical Sciences, Tehran, Iran.Debora HenryBritish Columbia’s Research Institute for Children’s and Women’s Health. Vancouver, British Columbia, Canada.David P. SpeertBritish Columbia’s Research Institute for Children’s and Women’s Health. Vancouver, British Columbia, Canada
Division of Infectious Diseases, Department of Pediatrics, Faculty of Medicine, University of British
Columbia, Vancouver, Canada.Journal Article20030401Sixty four Iranian patients with cystic fibrosis (CF) were studied for colonization with Pseudomonas<br />aeruginosa. The patient’s age ranged between 2 months to 18 years old. Twenty one patients were<br />colonized, 15 with non-mucoid and 6 with mucoid strains of P. aeruginosa. The colonization rate<br />increased with age and the mucoid phenotype was only recovered from the older patients. All mucoid<br />strains came from patients with respiratory disease whereas most of the non-mucoid isolates (n=13)<br />were from patients with gastrointestinal disorder. The antibiograms of the isolates showed 100% sensitivity<br />to Imipenem and Collistin followed by Ciprofloxacin (90.5%), Ceftazidime, and Tobramycin (85.7%),<br />Amikacin, Piperacillin and Tazobactam-Piperacillin (81%), Ticarcillin (76%), Gentamycin (62%),<br />Mezlocillin (52.4%) and Carbenicillin (43%). The MICs for Ceftazidime, Gentamycin and Tobramycin<br />agreed with the disk test results. However, MIC determination for Amikacin showed a 100% sensitivity<br />compared to the disk test where 81% sensitivity was observed. The discrepancy may be due to the fact<br />that over 20% of the isolates had borderline MIC values for Amikacin. The genomic fingerprinting of the<br />21 isolates as well as the non-mucoid revertants ofthe mucoid strains was carried out by RAPD-PCR<br />using primer 272 which was previously used for typing P. aeruginosa isolates from CF patient’s. Thirteen<br />genotypes were found among the 21 isolates. One fingerprint (A) was found in 6 patients and another<br />(B) was shared by 2 patients, all from the same health center. The idea of the hospital as environmental<br />source or cross infection between patients cannot be ruled out.<br /><br />National Institute of Genetic Engineering and Biotechnology of IranIranian Journal of Biotechnology1728-30431220030401Comparison of T7- and Lac-Based Systems for the Periplasmic Expression of Human Granulocyte Macrophage Colony Stimulating Factor in Escherichia coli1011086874ENShirin BorjalilooNational Research Center for Genetic Engineering and Biotechnology, P.O. Box: 14155-6343, Tehran, Iran.Alireza ZomorodipourNational Research Center for Genetic Engineering and Biotechnology, P.O. Box: 14155-6343, Tehran, Iran.0000-0003-0671-4779Bagher YakhchaliNational Research Center for Genetic Engineering and Biotechnology, P.O. Box: 14155-6343, Tehran, Iran.0000-0003-4031-877xSharareh ShojaiNational Research Center for Genetic Engineering and Biotechnology, P.O. Box: 14155-6343, Tehran, Iran.Journal Article20030401With the aim of the production of recombinant human granulocyte macrophage colony stimulating factor (rhGM-CSF) in the periplasmic space of Escherichia coli, the expression of hGM-CSF cDNA was examined<br />under the regulation of a T7-based as well as a lac-based expression systems. For the efficient expression of hGM-CSF cDNA, the first five codons at the N-terminal were altered based on the E. coli major codon usage. The hGM-CSF cDNA, fused to pelB signal sequence, was expressed using the two inducible promoters. The expression analysis of the 2 recombinant plasmids were performed in the BL21(DE3) and TG1 strains of E. coli, respectively. After induction with 1mM isopropyl-ß-DThiogalactopyranoside (IPTG) the recombinant E.<br />coli with T7 promoter produced hGM-CSF more efficiently than did the lac promoter. Under inducing conditions<br />both of the recombinant bacteria allowed successful secretion of hGM-CSF into the periplasmic space. The optimal temperature for the over-expression of the recombinant protein under the T7-based system was 30°C and that of the lac regulated system was 28°C. The optimization of growth condition for the recombinant bacteria, produced in this work, provides mean for studying the function of environmental as well as genetic factors on the overexpression of recombinant proteins in the periplasmic space of E. coli.National Institute of Genetic Engineering and Biotechnology of IranIranian Journal of Biotechnology1728-30431220030401Expression of Recombinant 3-Beta Hydroxysteroid Dehydrogenase Protein in E. coli1091146888ENSadeq VallianDivision of Genetics, Department of Biology, Faculty of Science, Isfahan University. Isfahan, I.R. IranFarzin FarzanehMolecular Medicine Department, King´s College School of Medicine and Dentistry, King´s College, London, UK.Journal Article200304013-beta hydroxysteroid dehydrogenase (3BHSD) is secreted by the cortex of adrenal gland functioning in<br />stress conditions. The gene encoding the 3-beta hydroxysteroid dehydrogenase protein was PCRamplified from a λgt11 cDNA library using specific primers. The amplified PCR product was then cloned into pGEX-4T-1 expression vector under Ptac promoter and the expression of the enzyme was examined in E. coli (BL21). Upon optimization of the expression condition, the enzyme was produced as a glutathione S-transferase (GST) fusion protein, which was purified by affinity chromatography using glutathione sepharose column. The GST part was then removed by selective proteolytic digestion with thrombin. The purified recombinant enzyme could be used in construction of diagnostic kits for screening the patients with premature ovarian failure (POF) for the presence of autoantibodies against 3BHSD as an important molecular target.National Institute of Genetic Engineering and Biotechnology of IranIranian Journal of Biotechnology1728-30431220030401Study on Nitrification and Denitrification of High Nitrogen and COD Load Wastewater in Moving Bed Biofilm Reactor1151206896ENS JonoudChemical and Petroleum Engineering Department, Sharif University of Technology, Tehran, Iran.M VosoughiChemical and Petroleum Engineering Department, Sharif University of Technology, Tehran, Iran.N Khalili DaylamiChemical and Petroleum Engineering Department, Sharif University of Technology, Tehran, Iran.Journal Article20030401Removing nitrogen, as one of the most common and abundant pollutant of ground and surface waters is<br />very important. For this purpose, biological nitrification and denitrification as the most economical method should be investigated. Feasibility of high load (Chemical Oxygen Demand) COD (1000-2000 mg/l) and NH4 (1000-2000 mg/l) wastewater treatment, at different Hydraulic Retention Times (HRTs) was studied in two 9-lit anaerobic-aerobic system in pre-denitrification mode. Moving Bed Biofilm Reactor (MBBR) is a new system, having all the advantages of activated sludge, fluidized bed and fixed bed processes, without disadvantages of each system, that the biofilm production takes place on the packings, moving along the height of the reactor. From the experiments carried out in this system, result, showed higher ammonia removals take place at higher ammonia and lower organic loads. Denitrification increases at higher nitrification rates because of the<br />increasing effect of NO3 - entering the anaerobic reactor. In spite of the fact that nitrifying bacteria are more sensitive than COD and NO3 - removing bacteria, after toxic shock by phenol as organic source,<br />nitrification rate increases and COD removal decreases according to the damaging effect of phenol on COD removing bacteria. Total COD removal during the study varies between 80-100%, this value changes to 30-80% for ammonia and 30-80% for ammonia and 40-90% for nitrate.National Institute of Genetic Engineering and Biotechnology of IranIranian Journal of Biotechnology1728-30431220030401Isolation of a Dibenzothiophene Desulfurizing Bacterium from Soil of Tabriz Oil Refinery1211246893ENFiroozeh Khadem HaghighatBiotechnology Center of Iranian Research Organization for Science and Technology, No.71, Forsat St., Enghelab Ave., Tehran, Iran. Biology Department, Faculty of Science, Shahid Beheshti University, Evin, Tehran, Iran.Fereshteh EftekharBiology Department, Faculty of Science, Shahid Beheshti University, Evin, Tehran, Iran.Mahnaz MazaheriBiotechnology Center of Iranian Research Organization for Science and Technology, No.71, Forsat St., Enghelab Ave., Tehran, Iran.Journal Article20030401A dibenzothiophene (DBT) degrading bacterium, which utilizes DBT as the sole source of sulfur, was isolated from soil contaminated with crud oil collected from oil refinery of Tabriz (Iran). A convenient spectrophotometric assay (Gibbs’ assay) was used to determine the quantity of desulfurized product<br />(Hydroxybiphenyl). This isolate did not grow on DBT, dibenzothiophene sulfone (DBTO2), or 2-Hydroxybiphenyl (HBP) as sole carbon sources. Biodesulfurization activity was observed only in growing<br />cultures and depressed by free sulfate. The desulfurization trait was expressed at increasing levels during the exponential phase of growth and then declined in stationary-phase cells. This gram-positive, non-spore-forming, partially acid fast, polymorphic bacterium with the ability to desulfurize DBT or DBTO2 was identified as Rhodococcus sp. and designated strain FMF.