ORIGINAL_ARTICLE
Variability of the Cyclin-Dependent Kinase 2 Flexibility Without Significant Change in the Initial Conformation of the Protein or Its Environment; a Computational Study
Background: Protein flexibility, which has been referred as a dynamic behavior has various roles in proteins’ functions. Furthermore, for some developed tools in bioinformatics, such as protein-protein docking software, considering the protein flexibility, causes a higher degree of accuracy. Through undertaking the present work, we have accomplished the quantification plus analysis of the variations in the human Cyclin Dependent Kinase 2 (hCDK2) protein flexibility without affecting a significant change in its initial environment or the protein per se.Objectives: The main goal of the present research was to calculate variations in the flexibility for each residue of the hCDK2, analysis of their flexibility variations through clustering, and to investigate the functional aspects of the residues with high flexibility variations.Materials and Methods: Using Gromacs package (version 4.5.4), three independent molecular dynamics (MD) simulations of the hCDK2 protein (PDB ID: 1HCL) was accomplished with no significant changes in their initial environments, structures, or conformations, followed by Root Mean Square Fluctuations (RMSF) calculation of these MD trajectories. The amount of variations in these three curves of RMSF was calculated using two formulas.Results: More than 50% of the variation in the flexibility (the distance between the maximum and the minimum amount of the RMSF) was found at the region of Val-154. As well, there are other major flexibility fluctuations in other residues. These residues were mostly positioned in the vicinity of the functional residues. The subsequent works were done, as followed by clustering all hCDK2 residues into four groups considering the amount of their variability with respect to flexibility and theirposition in the RMSF curves.Conclusions: This work has introduced a new class of flexibility aspect of the proteins’ residues. It could also help designing and engineering proteins, with introducing a new dynamic aspect of hCDK2, and accordingly, for the other similar globular proteins. In addition, it could provide a better computational calculation of the protein flexibility, which is, especially important in the comparative studies of the proteins’ flexibility.
https://www.ijbiotech.com/article_81323_363a4c664563afcfe0f6166c93dda2c3.pdf
2016-06-01
1
12
10.15171/ijb.1419
Flexibility fluctuation
Human CDK2 (hCDK2) protein
Molecular Dynamics-Root Mean Square Fluctuation (MD-RMSF)
Molecular Dynamics Simulation
Protein flexibility
RMSF Standard Deviation (RMSF-SD)
Mohammad
Taghizadeh
mtaghizadeh@ut.ac.ir
1
Laboratory of Biophysics and Molecular Biology, Institute of Biochemistry and Biophysics, University of Tehran, Tehran, Iran
AUTHOR
Bahram
Goliaei
goliaei@ibb.ut.ac.ir
2
Laboratory of Biophysics and Molecular Biology, Institute of Biochemistry and Biophysics, University of Tehran, Tehran, Iran
LEAD_AUTHOR
Armin
Madadkar Sobhani
arminms@gmail.com
3
Institute of Biochemistry and Biophysics (IBB), Tehran University, Tehran, Iran.
AUTHOR
ORIGINAL_ARTICLE
Optimization of Carbon and Nitrogen Sources for Extracellular Polymeric Substances Production by Chryseobacterium indologenes MUT.2
Background: Bacterial Extracellular Polymeric Substances (EPS) are environmental friendly and versatile polymeric materials that are used in a wide range of industries such as: food, textile, cosmetics, and pharmaceuticals. To make the production process of the EPS cost-effective, improvements in the production yield is required which could be implemented through application of processes such as optimized culture conditions, and development of the strains with higher yield (e.g. through genetic manipulation), or using low-cost substrates. Objectives: In this work, the effects of carbon and nitrogen sources were studied in order to improve the EPS production by the submerged cultivation of Chryseobacterium indologenes MUT.2. Materials and Methods: The mesophilic microorganism Chryseobacterium indologenes MUT.2, was grown and maintained in the Luria Bertani agar. The initial basal medium contained: glucose (20 g.L-1), yeast extracts (5 g.L-1), K2HPO4 (6 g.L-1), NaH2PO4 (7 g.L-1), NH4CL (0.7 g.L-1), and MgSO4 (0.5 g.L-1). For evaluating the carbon and nitrogen sources’ effect on the fermentation performance, cultures were prepared in 500 mL flasks filled with 300 mL of the medium. The single-factor experiments based on statistics was employed to evaluate and optimize the carbon and nitrogen sources for EPS production in the liquid culture medium of Chryseobacterium indologenes MUT.2. Results: The preferred carbon-sources, sucrose and glucose, commonly gave the highest EPS production of 8.32 and 6.37 g.L-1, respectively, and the maximum EPS production of 8.87 g.L-1 was achieved when glutamic acid (5 g.L-1) was employed as the nitrogen source. Conclusions: In this work, the culture medium for production of EPS by Chryseobacterium indologenes MUT.2 was optimized. Compared to the basal culture medium in shake-flasks and stirred tank bioreactor, the use of optimized culture medium has resulted in a 53% and 73% increase in the EPS production, respectively.
https://www.ijbiotech.com/article_14132_0c9a98e8683b813ca91b5244080c4603.pdf
2016-06-01
13
18
10.15171/ijb.1266
Carbon source
Chryseobacterium indologenes
Extracellular polymeric substance
Medium composition
Nitrogen source
Stirred tank bioreactor
Mojtaba
khani
mj_khani67@yahoo.com
1
Department of Bioscience and Biotechnology, Malek Ashtar University, Tehran, Iran.
AUTHOR
Ali
Bahrami
a_bahrami@mut.ac.ir
2
Department of Bioscience and Biotechnology, Malek Ashtar University, Tehran, Iran.
LEAD_AUTHOR
Asma
Chegeni
asma_chegeni@yahoo.com
3
Department of Bioscience and Biotechnology, Malek Ashtar University, Tehran, Iran.
AUTHOR
Mohammad Davoud
Ghafari
m.davoudghafari@gmail.com
4
Young Researchers and Elites Club, North Tehran Branch, Islamic Azad University, Tehran, Iran.
AUTHOR
Ali
Mansouran zadeh
a_bahrami@yahoo.com
5
Department of Bioscience and Biotechnology, Malek Ashtar University, Tehran, Iran.
AUTHOR
1. Khani M, Bahrami A, Ghafari MD. Optimization of operating parameters for anti-corrosive biopolymer production by Chryseobacterium Indologenes MUT. 2 using central composite design methodology. J Taiwan Inst Chem Eng. 2015. DOI: 10.1016/j.jtice.2015.09.016
1
2. Alves VD, Freitas F, Torres CA, Cruz M, Marques R, Grandfils C, et al. Rheological and morphological characterization of the culture broth during exopolysaccharide production by Enterobacter sp. Carbohydr Polym. 2009. DOI: 10.1016/j.carbpo l.2009.09.006
2
3. Freitas F, Alves VD, Reis MA. Advances in bacterial exopolysaccharides: from production to biotechnological applications. Trends Biotechnol Res. 2011;29(8):388-398. DOI: 10.1016/j.tibtech.2011.03.008
3
4. Suresh Kumar A, Mody K, Jha B. Bacterial exopolysaccharides–a perception. J Basic Microbiol. 2007;47(2):103-117. DOI: 10.1002/jobm.200610203
4
5. Kim SW, Xu CP, Hwang HJ, Choi JW, Kim CW, Yun JW. Production and Characterization of Exopolysaccharides from an Enthomopathogenic Fungus Cordycepsmilitaris NG3. Biotechnol Prog. 2003;19(2):428-435. DOI: 10.1021/bp 025644k
5
6. Lopez JC, Pérez JS, Sevilla JF, Fernandez FA, Grima EM, Chisti Y. Production of lovastatin by Aspergillus terreus: effects of the C: N ratio and the principal nutrients on growth and metabolite production. Enzyme Microb Technol. 2003;33(2):270-277. DOI: 10.1016/S0141-0229(03)00130-3
6
7. Görke B, Stülke J. Carbon catabolite repression in bacteria: many ways to make the most out of nutrients. Nat Rev Microbiol. 2008;6(8):613-624. DOI: 10.1038/nrmicro1932
7
8. Ghafari M, Bahrami A, Rasooli I, Arabian D, Ghafari F. Bacterial exopolymeric inhibition of carbon steel corrosion. Int Biodeterior Biodegradation. 2013;80:29-33. DOI: 10.1016/j.ibiod.2013.02. 007
8
9. Galindo E, Salcedo G, Ramírez ME. Preservation of Xanthomonas campestris on agar slopes: effects on xanthan production. Appl Microbiol Biotechnol. 1994;40(5):634-637. DOI: 10.1007/BF00173320
9
10. Garcýa-Ochoa F, Santos V, Casas J, Gomez E. Xanthan gum: production, recovery, and properties. Biotechnol Adv. 2000;18(7):549-579. DOI: 10.1016/S0734-9750(00)00050-1
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11. Büchs J. Introduction to advantages and problems of shaken cultures. Biochem Eng J. 2001;7(2):91-98. DOI: 10.1016/S1369-703X(00)00106-6
11
12. Suresh S, Srivastava V, Mishra I. Critical analysis of engineering aspects of shaken flask bioreactors. Crit Rev Biotechnoly. 2009;29(4):255-278. DOI: 10.3109/07388550903062314
12
13. Garcia-Ochoa F, Gomez E. Bioreactor scale-up and oxygen transfer rate in microbial processes: an overview. Biotechnol Adv. 2009;27(2):153-176. DOI: 10.1016/j.biotechadv.2008. 10.006
13
14. Reyes C, Peña C, Galindo E. Reproducing shake flasks performance in stirred fermentors: production of alginates by Azotobacter vinelandii. J Biotechnol. 2003;105(1):189-198. DOI: 10.1016/ S0168-1656(03)00186-X
14
15. Kornmann H, Duboc P, Marison I, von Stockar U. Influence of nutritional factors on the nature, yield, and composition of exopolysaccharides produced by Gluconacetobacter xylinus I-2281. Appl Environ Microbiol. 2003;69(10):6091-6098. DOI: 10.1128/AEM.69.10.6091-6098.2003
15
16. Bueno SM, Garcia-Cruz CH. Optimization of polysaccharides production by bacteria isolated from soil. Braz J Microbiol. 2006;37(3):296-301. DOI: 10.1590/S1517-838220060003 00018
16
17. Zhang J, Dong YC, Fan LL, Jiao ZH, Chen QH. Optimization of culture medium compositions for gellan gum production by a halobacterium Sphingomonas paucimobilis. Carbohydr Polym. 2015;115:694-700. DOI: 10.1016/j.carbpol.2014.09. 029
17
18. Qiang L, Yumei L, Sheng H, Yingzi L, Dongxue S, Dake H, et al. Optimization of fermentation conditions and properties of an exopolysaccharide from Klebsiella sp. H-207 and application in adsorption of hexavalent chromium. PLoS ONE. 2013;8(1):e53542. DOI: 10.1371/journal.pone.0053542
18
19. Jathore NR, Bule MV, Tilay AV, Annapure US. Microbial levan from Pseudomonas fluorescens: Characterization and medium optimization for enhanced production. Food Sci Biotechnol. 2012;21(4):1045-1053. DOI: 10.1007/s10068-012-0136-8
19
20. Silvi S, Barghini P, Aquilanti A, Juarez-Jimenez B, Fenice M. Physiologic and metabolic characterization of a new marine isolate (BM39) of Pantoea sp. producing high levels of exopolysaccharide. Microb Cell Fact. 2013;12(1):10. DOI: 10.1186/1475-2859-12-10
20
21. Bounaix M-S, Gabriel Vr, Morel S, Robert H, Rabier P, Remaud-Sime´on M, et al. Biodiversity of exopolysaccharides produced from sucrose by sourdough lactic acid bacteria. J Agric Food Chem. 2009;57(22):10889-10897. DOI: 10. 1021/jf902068t
21
ORIGINAL_ARTICLE
VIT-CMJ2: Endophyte of Agaricus bisporus in Production of Bioactive Compounds
Background: Agaricus bisporus is an edible basidiomycete fungus. Both the body and the mycelium contain compounds comprising a wide range of antimicrobial molecules, contributing in improvement of immunity and tumor-retardation. Objectives: The presence of endophytes capable of producing bioactive compounds was investigated in Agaricus bisporus. Materials and Methods: Endophytes from Agaricus bisporus was isolated on LB agar. The obtained isolates were characterized morphologically and biochemically. Further 16S rRNA sequencing was implemented for molecular analysis of isolates. The isolate was mass produced and the bioactive compounds were extracted using ethyl acetate, chloroform and hexane. Agar well diffusion method was carried out to seek the potential of any antimicrobial activity of the crude bioactive compounds against known pathogens. GC-MS and FT-IR analysis were performed for the identification of bioactive compounds. Results: VIT-CMJ2 was identified as Enterobacter sp. as revealed by 16S rRNA sequencing. Chloroform extract of VIT-CMJ2 showed a maximum zone of inhibition of 19 mm against Salmonella typhi followed by hexane and ethyl acetate extracts. The GC-MS analysis revealed the presence of several bioactive compounds having effective antimicrobial activity like butyl ester, Behenicalcohol, S , S-dioxide derivatives and some others which were later confirmed by FT-IR spectral stretches. Conclusions: The present study shows the insight on the way endophytes interact with Agaricus bisporus; thereby improving the nutritional profile.
https://www.ijbiotech.com/article_14133_21a0dba1c214d1170ed71ded6ef5db69.pdf
2016-06-01
19
24
10.15171/ijb.1287
Antibacterial Activity
Behenic alcohol
Button mushroom
Butyl ester
Endophytes
FT-IR
GC-MS
16S rRNA
Chandan
Gautam
ckg310@gmail.com
1
Department of Biomolecules Lab, School of Bio Sciences and Technology, VIT University, Vellore, India
AUTHOR
Mukund
Madhav
mukund.madhav51@gmail.com
2
Department of Biomolecules Lab, School of Bio Sciences and Technology, VIT University, Vellore, India
AUTHOR
Astha
Sinha
astha21.vit@gmail.com
3
Department of Biomolecules Lab, School of Bio Sciences and Technology, VIT University, Vellore, India
AUTHOR
William
Osborne
jabez.vit@gmail.com
4
Department of Biomolecules Lab, School of Bio Sciences and Technology, VIT University, Vellore, India
LEAD_AUTHOR
1. Sanmee R, Dell B, Lumyong P, Izumori K, Lumyong S. Nutritive value of popular wild edible mushrooms from northern Thailand. Food Chem. 2003;82:527-532. DOI: 10.1016/ S0308-8146(02)00595-2
1
2. Chang ST. Global impact of edible and medicinal mushrooms on human welfare in 21st century: Nongreen revolution. Int J Med Mushrooms. 1999;1:1-7. DOI: 10.1615/IntJMedMushrooms.v1. i1.10
2
3. Aida FMNA, Shuhaimi M, Yazid M, Maaruf AG. Mushroom as a potential source of prebiotics: a review. Trends Food Sci Tech. 2009;20:567-575. DOI: 10. 1016/j.tifs.2009.07.007
3
4. Xu X, Yan H, Chen J, Zhang X. Bioactive proteins from mushrooms. Biotechnol Adv. 2011;29:667-674. DOI: 10.1016/j. biotechadv.2011.05.003
4
5. Mattilaa PA, Ronkainen R, Toivob J, Piironen V. Sterol and vitamin D2 contents in some wild and cultivated mushrooms. Food Chem. 2002;76:293-298. DOI: 10.1016/S0308-8146 (01)00275-8
5
6. Bao, X, Wang X, Dong Q, Fang J, Li X. Structural features of immunologically active polysaccharides from Ganoderma lucidum. Phytochemistry 2002;59:175-181. DOI: 10.1016/S0031-9422(01)00450-2
6
7. Zarenejad F, Yakhchali B, Rasooli I. Evaluation of indigenous potent mushroom growth promoting bacteria (MGPB) on Agaricus bisporus production. World J Microbiol Biotechnol. 2012;28:99-104. DOI: 10.1007/s11274-011-0796-1
7
8. Cho Y, Kim, J, Crowley DE, Cho B. Growth promotion of the edible fungus Pleurotus ostreatus by fluorescent pseudomonads. FEMS Microbiol Lett. 2003;218:271-276. DOI: 10.1016/ S0378-1097(02)01144-8
8
9. Young L, Chu J, Hameed A, Young C. Cultivable mushroom growth-promoting bacteria and their impact on Agaricus blazei productivity. Pesq agropec bras Brasília. 2013;48:636-644. DOI: org/10.1590/S0100-204X2013000600009
9
10. El Enshasy HA, Hatti-Kaul R. Mushroom immunomodulators: unique molecules with unlimited applications. Trends Biotechnol. 2013;13:668-677. DOI: 10.1016/j.tibtech.2013.09.003
10
11. Vinodhkumar T, Maithili SS, Ramanathan G, Sudhakar .Antibacterial properties of secondary metabolites from endophytic marine algal bacterial population against chicken meat microbial pathogen. Int J Curr Sci. 2013;6:133-139.
11
12. Joseph B, Priya RM, Helen PAM, Sujatha S. Bio-active compounds in essential oil and its effects of antimicrobial, cytotoxic activity from the Psidiumguajava (L.) leaf. J Adv Biotechnol. 2010;9:10-14.
12
13. Yuvaraj N, Kanmani P, Satishkumar R, Paari KA, Pattukumar V, Arul V. Extraction, purification and partial characterization of Cladophor aglomerata against multidrug resistant human pathogen Acinetobacter baumannii and fish. World J Fish Mar Sci. 2011;3(1):51-57.
13
14. Buchholz A, Takors R, Wandrey C. Quantification of intracellular metabolites in Escherichia coli K12 using liquid chromatographic-electrospray ionization tandem mass spectrometric techniques. Anal Biochem. 2001;295:129-137. DOI: 10.1006/abio.2001.5183
14
15. Fox A. Carbohydrate profiling of bacteria by gas chromatography-mass spectrometry and their trace detection in complex matrices by gas chromatography-tandem mass spectrometry. J Chromatogr A. 1999;843:287-300. DOI: 10.1016/S0021-9673(98)00884-X
15
16. Saravanan V, Osborne J, Madhaiyan M, Mathew L, et al. Zinc metal solubilization by Gluconacetobacter diazotrophicus and induction of pleomorphic cells. J Microbiol Biotechnol. 2007;17(9):1477-1482.
16
17. Chen T, Chen Z, Ma GH, Du BH, Shen B, Ding YQ, Xu K. Diversity and potential application of endophytic bacteria in ginger. Genet Mol Res. 2014;13(3):4918-4931. DOI: 10.4238/ 2014.July.4
17
18. Mbai FN, Magiri EN, Matiru VN, Nganga J, Nyambati VCS. Isolation and characterisation of bacterial root endophytes with potential to enhance plant growth from kenyan basmati rice. Am Int J Contemp Res. 2013;3(4):25-40.
18
19. Guo B, Wang Y, SunX,Tang K. Bioactive natural products from endophytes: A review. Appl MicrobiolBiotechnol. 2008;44(2):136-142. DOI: 10.1134/S0003683808020026
19
20. Merkl R, Iveta H, Vladimir F, Jan S. Antimicrobial and antioxidant properties of phenolic acids alkyl esters. Czech J Food Sci. 2010;28(4):275-279.
20
21. Kokate CK, Regelson W. Review of the biology of Quercetin and related bioflavanoids. Food Chem Toxicol. 1995;33:1061-1080.
21
ORIGINAL_ARTICLE
Green Extracellular Synthesis of the Silver Nanoparticles Using Thermophilic Bacillus Sp. AZ1 and its Antimicrobial Activity Against Several Human Pathogenetic Bacteria
Background: Silver nanoparticles (AgNPs) are among the most effective antimicrobial agents that are used in the medicine and pharmaceutics. During the past decades, metal nanoparticles synthesis through application of the biological methods has increasingly been used, as the biologically synthesized particles are mostly non-toxic as well as effective. Objectives: The main goal for undertaking the present investigation was to evaluate the extracellular synthesis of the AgNPs by a native thermophilic Bacillus Sp. AZ1 that was isolated from a hot spring in Ardebil province. Subsequently the antimicrobial potentials of the nanoparticle was evaluated against several human pathogenic organisms. Materials and Methods: The biosynthesized AgNPs were confirmed visually by appearance of a dark brown color formation in the mixture as well as silver surface plasmon resonance band by using UV-Visible spectroscopy. The AgNPs were further characterized by SEM, EDX and TEM. The antimicrobial activity of the AgNPs was investigated using Salmonella typhi, Escherichia coli, Staphylococcus epidermis, and Staphylococcus aureus, by applying disk diffusion method. Results: Identification of the strain AZ1 by the 16S rRNA sequence analysis showed 99% sequence homology between this strain and B. licheniformis. The obtained UV-Visible spectrum of the aqueous medium containing silver ion, showed a peak at 425 nm which indicates a correspondence to the plasmon absorbance of the silver nanoparticles. The biosynthesized AgNPs were found to be in the size range of ~7-31 nm with spherical the shape. Studies regarding the antibacterial effect of the particles showed the highest inhibitory effect against the two strains; E. coli, and S. typhi, respectively. Conclusions: Our study presents a simple green synthesis process for the production of an extracellular nanoparticles which is environmental friendly. Biosynthesis of the AgNPs by a thermophilic bacillus from the hot spring (Qeynarjeh, Ardebil) in Iran with the highest similarity to Bacillus licheniformis is reported for the first time.
https://www.ijbiotech.com/article_14130_f84bd584aa6275924b43faeb389471d6.pdf
2016-06-01
25
32
10.15171/ijb.1259
Antimicrobial agents
Biosynthesis
Nanoparticles
16S rRNA
Ali
Deljou
alideljou@yahoo.com
1
Department of Biotechnology, Faculty of Agriculture, Bu-Ali Sina University, Hamedan, Iran
LEAD_AUTHOR
samad
Goudarzi
smdgoudarzi@gmail.com
2
Buali sina
LEAD_AUTHOR
1. Bruinsma N, Kristinsson KG, Bronzwaer S, Schrijnemakers P, Degener J, Tiemersma E, et al. Trends of penicillin and erythromycin resistance among invasive Streptococcus pneumoniae in Europe. J Antimicrob Chemother. 2004;54(6):1050-1045. DOI: 10.1093/jac/dkh458
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3. Threlfall EJ. Antimicrobial drug resistance in Salmonella: problems and perspectives in food-and water-borne infections. FEMS Microbiol Rev. 2002;26(2):148-141. DOI: 10.1111/j.1574-6976. 2002.tb00606.x
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4. Tenover FC. Mechanisms of antimicrobial resistance in bacteria. Am J Med. 2006;119(6):10-3. DOI:10.1016/j.amjmed.2006.03. 03.011
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5. Sukdeb P, Kyung TY, Myong SJ. Does the antibacterial activity of silver nanoparticles depend on the shape of the nanoparticle? A study of the gram-negative bacterium E. coli. Appl Environ Microbiol. 2007;73(6):1720-1712. DOI: 10.1128/ AEM.02218-06
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6. Chudasama B, Vala A K, Andhariya N, Mehta RV, Upadhyay R V. Highly bacterial resistant silver nanoparticles: synthesis and antibacterial activities. J Nanopart Res. 2010;12(5):1685-1677. DOI: 10.1007/s11051-009-9845-1
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8. Rai M K , et al. Silver nanoparticles: the powerful nanoweapon against multidrug-resistant bacteria. J Appl Microbiol. 2012;112(5):852-841. DOI: 10.1111/j.1365-2672.2012.05253.x
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9. Kim JS, Kuk E, Yu KN, Kim JH, Park SJ, Lee HJ, et al. Antimicrobial effects of silver nanoparticles. Nanomedicine: NBM. 2007;3(1):101-95. DOI: 10.1016/j.nano.2006.12.001
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10. Prabhu S, Eldho K. Silver nanoparticles: mechanism of antimicrobial action, synthesis, medical Poulose applications, and toxicity effects. Int Nano Lett. 2012;2(1):10-1. DOI: 10.1186/2228-5326-2-32
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11. Singhal G, Bhavesh R, Kasariya K, Sharma AR, Singh RP. Biosynthesis of silver nanoparticles using Ocimum sanctum (Tulsi) leaf extract and screening its antimicrobial activity. J Nanopart Res. 2011;13(7):2988-2981. DOI: 10.1007/s11051-010-0193-y
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12. Iravani Korbekandi H, Mirmohammadi SV, Zolfaghari SB. Synthesis of silver nanoparticles: chemical, physical and biological methods. Res Pharm Sci. 2014;9(6):406-385.
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13. Kalishwaralal K, Deepak V, Ramkumarpandian S, Nellaiah H, Sangiliyandi G. Extracellular biosynthesis of silver nanoparticles by the culture supernatant of Bacillus licheniformis. Mater Lett. 2008;62(29):4413-4411. DOI: 10.1016/j.matlet. 2008.06.051
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14. Vilchis-Nestor AR, Sánchez-Mendieta V, Camacho-López MA, Gómez-Espinosa RM, Camacho-López MA, Arenas-Alatorre J A. Solventless synthesis and optical properties of Au and Ag nanoparticles using Camellia sinensis extract. Mater Lett. 2008;62(17):3105-3103. DOI: 10.1016/j.matlet.2008.01.138
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15. Gholami Shabani M, Akbarzadeh A, Norouzian D, Amini A, Gholami-Shabani Z, ImaniA, Chiani M, Riazi Gh, Shams-Ghahfarokhi M, Razzaghi M. Antimicrobial Activity and Physical Characterization of Silver Nanoparticles Green Synthesized using Nitrate Reductase from Fusarium oxysporum. Appl Biochem Biotechnol. 2014;172(8):4098-4084. DOI: 10.1007/s12010-014-0809-2
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18. Samadi N, Golkaran D, Eslamifar A, Jamalifar, H, Fazeli MR, Mohseni FA. Intra/extracellular Biosynthesis of Silver Nanoparticles by an Autochthonous Strain of Proteus mirabilis Isolated from Photographic Waste. J Biomed Nanotechnol. 2009;5(3):253-247. DOI: 10.1166/jbn.2009.1029
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31. Sunkar S, Nachiyar CV. Biogenesis of antibacterial silver nanoparticles using the endophytic bacterium Bacillus cereus isolated from Garcinia xanthochymus. Asian Pac J Trop Biomed. 2012;2(12):959-953. DOI: 10.1016/S2221-1691 (13)60006-4
31
32. Saravanan M, Anil KV, Sisir KB. Rapid biosynthesis of silver nanoparticles from Bacillus megaterium (NCIM 2326) and their antibacterial activity on multi drug resistant clinical pathogens. Colloids Surf B. 2011;88(1):331-332. DOI: 10.1016/j.colsurfb.2011.07.009
32
33. Durán N, Marcato PD, Durán M, Yadav A, Gade A, Rai M. Mechanistic aspects in the biogenic synthesis of extracellular metal nanoparticles by peptides, bacteria, fungi, and plants. Appl Microbiol Biotechnol. 2011;90(5):1624-1609. DOI: 10.1007/s00253-011-3249-8
33
34. Chudasama, Bhupendra, Vala AK, Andhariya N, Mehta RV, Upadhyay RV. Highly bacterial resistant silver nanoparticles: synthesis and antibacterial activities. J Nanopart Res. 2010;12(5): 1685-1677. DOI: 10.1007/s11051-009-9845-1
34
35. Ramgopal M, Saisushma CH, Abobaker M. Alhasin. A facile green synthesis of silver nanoparticles using soap nuts. Res J Microbiol. 2011;6(5):438-432. DOI: 10.3923/jm.2011.332.438
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36. Prakash A, Sharma S, Ahmad N, Ghosh A, Sinha P. Synthesis of AgNps By Bacillus cereus bacteria and their antimicrobial potential. J Biomater Nanobiotechnol. 2011;2(2):162-156. DOI: 10.4336/jbnb.2022.22020
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37. Priyadarshini S, Gopinath V, Priyadharsshini NM, MubarakAli D, Velusamy P. Synthesis of anisotropic silver nanoparticles using novel strain, Bacillus flexus and its biomedical application. Colloids Surf B. 2013;102(1):237-232. DOI: 10.1016/j.colsurfb. 2012.08.018
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38. Mahendra R, Yadav A, Aniket Gade. Silver nanoparticles as a new generation of antimicrobials. Biotechnol Adv. 2009;27(1):83-76. DOI: 10.1016/j.biotechadv.2008.09.002
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39. Li WR, Xie XB, Shi QS, Zeng HY, You-Sheng OY, Chen YB. Antibacterial activity and mechanism of silver nanoparticles on E. coli. Appl Microbiol Biotechnol. 2010;85(4):1122-1115. DOI: 10.1007/s00253-009-2159-5
39
40. Marambio J, Hoek MVC, Hoek MVE. A review of the antibacterial effects of silver nanomaterials and potential implications for human health and the environment. J Nanopart Res. 2010;12(5):1551-1531. DOI: 10.1007/s11051-010-9900-y
40
ORIGINAL_ARTICLE
Optimization of RGD-modified Nano-liposomes Encapsulating Eptifibatide
Background: Eptifibatide (Integrilin) is an intravenous (IV) peptide drug that selectively inhibits ligand binding to the platelet GP IIb/IIIa receptor. It is an efficient peptide drug, however has a short half-life. Therefore, antithrombotic agents like eptifibatide are required to become improved with a protected and targeted delivery system such as using nano-liposomes to the site of thrombus. Objectives: The goal in the present report was to optimize encapsulation efficiency of the eptifibatide into Arg-Gly-Asp (RGD)-modified nano-liposomes (RMNL). As well, it was intended to evaluate the effect of sodium lauryl sulfate (SLS) on drug release. Materials and Methods: The effect of five independent variables including number of freeze/thawing cycles, concentration of eptifibatide, 1,2-distearoyl-sn-glycero-3-phosphocholine (DSPC), cholesterol, and dipalmitoyl-GRGDSPA peptide on drug entrapment efficiency (DEE) was investigated using response surface methodology (RSM). The effect of different concentrations of SLS on encapsulation and drug release from RMNL was also investigated. The size and morphology of RMNL were characterized using transmission electron microscopy (TEM). Results: The maximum DEE (38%) was obtained with 7 freeze/thawing cycles, 3.65 mmoL eptifibatide, 7 mM DSPC, 3 mM cholesterol, and 1 mM dipalmitoyl- GRGDSPA peptide. SLS has significantly increased the drug release from RMNL, although its effect on encapsulation efficiency was not significant. Conclusions: The optimization of the formulations for valuable and expensive peptide drugs is essential to have the maximum encapsulation efficiency and the minimum experiments.
https://www.ijbiotech.com/article_14135_82926dc54152825191a76d6b552ab2e7.pdf
2016-06-01
33
40
10.15171/ijb.1399
Eptifibatide
RGD-modified nano-liposomes (RMNL)
RMNL encapsulated eptifibatide
Hassan
Bardania
hbardania@gmail.com
1
Department of Nanobiotechnology, Faculty of Biological Sciences, Tarbiat Modares University, Tehran, Iran
AUTHOR
Seyed Abbas
Shojaosadati
shoja_sa@modares.ac.ir
2
Department of Biotechnology Group Chemical Engineering, Tarbiat Modares University, Tehran, Iran
LEAD_AUTHOR
Farzad
Kobarfard
kobarfard@sbmu.ac.ir
3
Department of Medical Chemistry, Faculty of Pharmacy, Shahid Beheshti University of Medical Sciences, Tehran, Iran
AUTHOR
Farid
Dorkoosh
dorkoosh@tums.ac.ir
4
Department of Pharmaceutics, School of Pharmacy, Tehran University of Medical Sciences, Tehran, Iran
AUTHOR
1. Sanz J, Fayad ZA. Imaging of atherosclerotic cardiovascular disease. Nature 2008;451:953-957. DOI: 10.1038/nature06 803
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2. Tcheng JE, O’Shea JC. Eptifibatide: a potent inhibitor of the platelet receptor integrin, glycoprotein IIb/IIIa. Expert Opin Investig Drugs. 1999;8(11):1893-1905. DOI: 10.1517/13543 784.8.11.1893
2
3. Addeo R, Faiola V, Guarrasi R, Montella L, Vincenzi B, Capasso E, et al. Liposomal pegylated doxorubicin plus vinorelbine combination as first-line chemotherapy for metastatic breast cancer in elderly women > or = 65 years of age. Cancer Chemother Pharmacol. 2008;62(2):285-292. DOI: 10.1007/s00280-007-0605-6
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4. Torchilin VP. Targeting of drugs and drug carriers within the cardiovascular system. Adv Drug Deliv Rev. 1995;17:75-101. DOI: 10.1016/0169-409X(95)00042-6
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5. Schiener M, Hossann M, Viola JR, Ortega-Gomez A, Weber C, Lauber K, et al. Nanomedicine-based strategies for treatment of atherosclerosis. Trends Mol Med. 2014;20(5):271-281. DOI: 10.1016/j.molmed.2013.12.001
5
6. Haller CA, Cui W, Wen J, Robson SC, Chaikof EL. Reconstitution of CD39 in liposomes amplifies nucleoside triphosphate diphosphohydrolase activity and restores thromboregulatory properties. J Vasc Surg. 2006;43(4):816-823. DOI: 10.1016/j.jvs.2005.11.057
6
7. Akbarzadeh A, Rezaei-Sadabady R, Davaran S, Joo SW, Zarghami N, Hanifehpour Y, et al. Liposome: classification, preparation, and applications. Nanoscale Res Lett. 2013;8(1):102. DOI: 10.1186/1556-276X-8-102
7
8. Srinivasan R, Marchant RE, Gupta AS. In vitro and in vivo platelet targeting by cyclic RGD-modified liposomes. J Biomed Mater Res A. 2010;93(3):1004-1015. DOI: 10.1002/ jbm.a.32549
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9. Vaidya B, Nayak MK, Dash D, Agrawal GP, Vyas SP. Development and characterization of site specific target sensitive liposomes for the delivery of thrombolytic agents. Int J Pharm. 2011;403:254-261. DOI: 10.1016/j.ijpharm.2010.10. 028
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10. Huang G, Zhou Z, Srinivasan R, Penn MS, Kottke-Marchant K, Marchant RE, et al. Affinity Manipulation of Surface-conjugated RGD-peptide to Modulate Binding of Liposomes to Activated Platelets. Biomaterials 2008;29(11):1676-1685. DOI: 10.1016/j.biomaterials.2007.12.015
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11. Lestini BJ, Sagnella SM, Xu Z, Shive MS, Richter NJ, Jayaseharan J, et al. Surface modification of liposomes for selective cell targeting in cardiovascular drug delivery. J Controlled Release. 2002;78:235-247. DOI: 10.1016/S0168-3659(01)00505-3
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12. Goodman SL, Cooper SL, Albrecht RM. Integrin receptors and platelet adhesion to synthetic surfaces. J Biomed Mater Res. 1993;27(5):683-695. DOI: 10.1002/jbm.820270516
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13. Fields GB, Noble RL. Solid phase peptide synthesis utilizing 9-fluorenylmethoxycarbonyl amino acids. Int J Pept Protein Res. 1990;35(3):161-214. DOI: 10.1111/j.1399-3011.1990. tb00939.x
13
14. Colletier JP, Chaize B, Winterhalter M, Fournier D. Protein encapsulation in liposomes: efficiency depends on interactions between protein and phospholipid bilayer. BMC Biotechnol. 2002;2:9. DOI: 10.1186/1472-6750-2-9
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15. Thiagarajan P, Kelly KL. Exposure of binding sites for vitronectin on platelets following stimulation. J Biol Chem. 1988;263(6):3035-3038.
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16. Schachter DM, Kohn J. A synthetic polymer matrix for the delayed or pulsatile release of water-soluble peptides. J Control Release. 2002;78(1):143-153. DOI: 10.1016/S0168-3659(01)00487-4
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17. Kouchakzadeh H, Shojaosadati SA, Maghsoudi A, Vasheghani Farahani E. Optimization of PEGylation conditions for BSA nanoparticles using response surface methodology. AAPS PharmSciTech. 2010;11:1206-1211. DOI: 10.1208/s12249-010-9487-8
17
18. Rocky-Salimi K, Hamidi-Esfahani Z, Abbasi S. Statistical optimization of arachidonic acid production by Mortierella alpina CBS 754.68 in submerged fermentation. Iran J Biotech. 2011;9(2):87-93.
18
19. Erik Westein UF, Christoph E. Hagemeyer and Karlheinz Peter. Destination Known: Targeted Drug Delivery in Atherosclerosis and Thrombosis. Drug Dev Res. 2013;74:460-471. DOI: 10.1002/ddr.21103
19
20. Zhang J, Ma G, Lv Z, Zhou Y, Wen C, Wu Y, et al. Targeted thrombolysis strategies for neuroprotective effect. Neural Regen Res. 2014;9(13):1316-1322. DOI: 10.4103/1673-5374.137580
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21. Vyas SP. Vaidya B. Targeted delivery of thrombolytic agents: role of integrin receptors. Expert Opin Drug Deliv. 2009;6(5):499-508. DOI: 10.1517/17425240902878002
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22. Gupta AS, Huang G, Lestini BJ, Sagnella S, Kottke-Marchant K, Marchant RE. RGD-modified liposomes targeted to activated platelets as a potential vascular drug delivery system. Thromb Haemost. 2005;93(1):106-114. DOI: 10.1160/TH04-06-0340
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23. Castile JD, Taylor KM. Factors affecting the size distribution of liposomes produced by freeze-thaw extrusion. Int J Pharm. 1999;188(1):87-95. DOI: 10.1016/S0378-5173(99)00207-0
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25. Holovati JL, Gyongyossy-Issa MI, Acker JP. Effects of trehalose-loaded liposomes on red blood cell response to freezing and post-thaw membrane quality. Cryobiology 2009;58(1):75-83. DOI: 10.1016/j.cryobiol.2008.11.002
25
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27. Ducat E BM, Lecomte F, Evrard B, Piel G. The experimental design as practical approach to develop and optimize a formulation of peptide-loaded liposomes. AAPS PharmSciTech. 2010;11(2):966-975. DOI: 10.1208/s12249-010-9463-3
27
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31
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32
ORIGINAL_ARTICLE
Osteogenic Differentiation and Mineralization on Compact Multilayer nHA-PCL Electrospun Scaffolds in a Perfusion Bioreactor
Background: Monolayer electrospun scaffolds have already been used in bone tissue engineering due to their high surface-to-volume ratio, interconnectivity, similarity to natural bone extracellular matrix (ECM), and simple production. Objectives: The aim of this study was to evaluate the dynamic culture effect on osteogenic differentiation and mineralizationi into a compact cellular multilayer nHA-PCL electrospun construct. The dynamic culture was compared with static culture. Materials and Methods: The calcium content, alkaline phosphatase (ALP) activity and cell viability were investigated on days 3 and 7. Results: When the dynamic culture compared to static culture, the mineralization and ALP activity were increased in dynamic culture. After 7 days, calcium contents were 41.24 and 20.44 mg.(cm3)-1, and also normalized ALP activity were 0.32 and 0.19 U.mg-1 in dynamic and static culture, respectively. Despite decreasing the cell viability until day 7, the scanning electron microscopy (SEM) results showed that, due to higher mineralization, a larger area of the construct was covered with calcium deposition in dynamic culture. Conclusions: The dynamic flow could improve ALP activity and mineralization into the compact cellular multilayer construct cultured in the perfusion bioreactor after 7 days. Fluid flow of media helped to facilitate the nutrients transportation into the construct and created uniform cellular construct with high mineralization. This construct can be applied for bone tissue engineering.
https://www.ijbiotech.com/article_14138_8bce3668921ec632085be849dbd5ffe5.pdf
2016-06-01
41
49
10.15171/ijb.1382
Mineralization
Electrospun scaffolds
Multilayer construct
Osteogenic differentiation
Perfusion bioreactor
Maliheh
Yaghoobi
m.yaghoobi@modares.ac.ir
1
Biomedical Engineering Group, Faculty of Chemical Engineering, Tarbiat Modares University, Tehran, Iran
AUTHOR
Sameereh
Hashemi-Najafabadi
s.hashemi@modares.ac.ir
2
Biomedical Engineering Group, Faculty of Chemical Engineering, Tarbiat Modares University, Tehran, Iran
LEAD_AUTHOR
Masoud
Soleimani
soleim_m@modares.ac.ir
3
Hematology Group, Faculty of Medical Sciences, Tarbiat Modares University, Tehran, Iran
AUTHOR
Ebrahim
Vasheghani-Farahani
evf@modares.ac.ir
4
Biomedical Engineering Group, Faculty of Chemical Engineering, Tarbiat Modares University, Tehran, Iran
AUTHOR
Seyyed Mohammad
Mousavi
mousavi_m@modares.ac.ir
5
Biotechnology Group, Faculty of Chemical Engineering, Tarbiat Modares University, Tehran, Iran
AUTHOR
1. Eap S, Ferrand A, Palomares CM, Hebraud A, Stoltz JF, Mainard D, et al. Electrospun nanofibrous 3D scaffold for bone tissue engineering. Biomed Mater Eng. 2012;22(1-3):137-141. DOI: 10.3233/bme-2012-0699
1
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2
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3
4. Rodrigues CAV, Fernandes TG, Diogo MM, da Silva CL, Cabral JMS. Stem cell cultivation in bioreators. Biotechnol Adv. 2011;29(6):815-829. DOI: 10.1016/j.biotechadv.2011.06.009
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6. Romagnoli C, Brandi ML. Adipose mesenchymal stem cells in the field of bone tissue engineering. World J Stem Cells. 2014;6(2):144-152. DOI: 10.4252/wjsc.v6.i2.144
6
7. Ajalloueian F, Tavanai H, Hilborn J, Donzel-Gargand O, et al. Emulsion electrospinning as an approach to fabricate PLGA/Chitosan nanofibers for biomedical applications. Biomed Res Int. 2014;2014:1-13. DOI: 10.1155/2014/475280
7
8. Llorens E, Armelin E, Pérez-Madrigal MDM, del Valle LJ, Alemán C, Puiggalí J. Nanomembranes and nanofibers from biodegradable conducting polymers. Polymers. 2013;5(3):1115-1157. DOI: 10.3390/polym5031115
8
9. Pillay V, Dott C, Choonara YE, Tyagi C, Tomar L, Kumar P, et al. A Review of the effect of processing variables on the fabrication of electrospun nanofibers for drug delivery applications. Biomed Res Int. 2013;2013:1-22.DOI:10.1155/2013/789289
9
10. Demir MM, Yilgor I, Yilgor E, Erman B. Electrospinning of polyurethane fibers. Polymer. 2002;43(11):3303-3309. DOI:10.1016/S0032-3861(02)00136-2
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11. Liang D, Hsiao BS, Chu B. Functional electrospun nanofibrous scaffolds for biomedical applications. Adv Drug Deliv Rev. 2007;59(14):1392-1412. DOI: 10.1016/j.addr.2007.04.02
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12. Yang F, Wolke JGC, Jansen JA. Biomimetic calcium phosphate coating on electrospun poly (e-caprolactone) scaffolds for bone tissue engineering. Chem Eng J. 2008;137(1):154-161. DOI:10.1016/j.cej.2007.07.076
12
13. Bao C, Chen W, Weir MD, Thein-Han W, Xu HHK. Effects of electrospun submicron fibers in calcium phosphate cement scaffold on mechanical properties and osteogenic differentiation of umbilical cord stem cells. Acta Biomater. 2011;7(11):4037-4044. DOI: 10.1016/j.actbio.2011.06.046
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14. Vasita R, Katti DS. Nanofibers and their applications in tissue engineering. Int. J Nanomedicine. 2006;1(1):15-30
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15. Ngiam M, Liao S, Patil AJ, Cheng Z, Chan CK, Ramakrishna S. The fabrication of nano-hydroxyapatite on PLGA and PLGA/collagen nanofibrous composite scaffolds and their effects in osteoblastic behavior for bone tissue engineering. Bone. 2009;45(1):4-16. DOI: 10.1016/j.bone.2009.03.674
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16. Swetha M, Sahithi K, Moorthi A, Srinivasan N, Ramasamy K, Selvamurugan N. Biocomposites containing natural polymers and hydroxyapatite for bone tissue engineering. Int. J Biol Macromol. 2010;47(1):1-4. DOI: 10.1016/j.ijbiomac.2010. 03.015
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17. Wang H, Li Y, Zuo Y, Li J, Ma S, Cheng L. Biocompatibility and osteogenesis of biomimetic nano-hydroxyapatite/polyamide composite scaffolds for bone tissue engineering. Biomaterials. 2007;28(22):3338-3348. DOI: 10.1016/j.biomaterials.2007.04.014
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18. Venugopal J, Prabhakaran MP, Zhang Y, Low S, Choon AT, Ramakrishna S. Biomimetic hydroxyapatite-containing composite nanofibrous substrates for bone tissue engineering. Philos Trans A Math Phys Eng Sci. 2010;368(1917):2065-2081. DOI: 10.1098/rsta.2010.0012
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19. Gaspar DA, Gomide V, Monteiro FJ. The role of perfusion bioreactors in bone tissue engineering. Biomatter. 2012;2(4):167-175. DOI: 10.4161/biom.22170
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20. Srouji S, Kizhner T, Suss-Tobi E, Livne E, Zussman E. 3-D Nanofibrous electrospun multilayered construct is an alternative ECM mimicking scaffold. J Mater Sci Mater Med. 2008;19(3):1249-1255. DOI: 10.1007/s10856-007-3218-z
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21. Madurantakam PA, Rodriguez IA, Garg K, McCool JM. Compression of multilayered composite electrospun scaffolds: a novel strategy to rapidly enhance mechanical properties and three dimensionality of bone scaffolds. Adv Mater Sci Eng. 2013. DOI: 10.1155/2013/561273
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23. Doustgani A, Vasheghani-Farahani E, Soleimani M, Hashemi-Najafabadi S. Optimizing the mechanical properties of electrospun polycaprolactone and nanohydroxyapatite composite nanofibers. Compos Part B Eng. 2012;43(4):1830-1836. DOI :10.1016/j.compositesb.2012.01.051
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24. Loh QL, Choong C. Three-dimensional scaffolds for tissue engineering applications: role of porosity and pore size. Tissue Eng Part B Rev. 2013;19(6):485-502. DOI: 10.1089/ten.TEB.2012.0437
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25. Owen TA, Aronow M, Shalhoub V, Barone LM, Wilming L, Tassinari MS, et al. Progressive development of the rat osteoblast phenotype in vitro: Reciprocal relationships in expression of genes associated with osteoblast proliferation and differentiation during formation of the bone extracellular matrix. J Cell Physiol. 1990;143(3):420-30. DOI: 10.1002/ jcp.1041430304
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27. Polo-Corrales L, Latorre-Esteves M, Ramirez-Vick JE. Scaffold design for bone regeneration. J Nanosci Nanotechnol. 2014;14(1):15-56. DOI: 10.1166/jnn.2014.9127
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29. Wang X, Ding B, Li B. Biomimetic electrospun nanofibrous structures for tissue engineering. Mater Today. 2013;16(6):229-241. DOI: 10.1016/j.mattod.2013.06.005
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30. Stops AJF, Heraty KB, Browne M, O’Brien FJ, McHugh PE. A prediction of cell differentiation and proliferation within a collagen–glycosaminoglycan scaffold subjected to mechanical strain and perfusive fluid flow. J Biomech. 2010;43(4):618-626. DOI: 10.1016/j.jbiomech.2009.10.037
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31. Yeatts AB, Fisher JP. Bone tissue engineering bioreactors: Dynamic culture and the influence of shear stress. Bone. 2011;48(2):171-181. DOI: 10.1016/j.bone.2010.09.138
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34. Rauh J, Milan F, Gunther KP, Stiehler M. Bioreactor systems for bone tissue engineering. Tissue Eng Part B Rev. 2011;17(4):263-280. DOI: 10.1089/ten.TEB.2010.0612
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41. Liao J, Guo X, Nelson D, Kasper FK, Mikos AG. Modulation of osteogenic properties of biodegradable polymer/extracellular matrix composite scaffolds generated with a flow perfusion bioreactor. Acta Biomater. 2010;6(7):2386-2393. DOI: 10.1016/j.actbio.2010.01.011
41
ORIGINAL_ARTICLE
Higher Expression Level and Lower Toxicity of Genetically Spliced Rotavirus NSP4 in Comparison to the Full-Length Protein in E. coli
Background: Rotavirus group A (RVA) is recognized as a major cause of severe gastroenteritis in children and new-born animals. Nonstructural protein 4 (NSP4) is responsible for the enterotoxic activity of these viruses in the villus epithelial cells. Amino acids 114-135 of NSP4 are known to form the diarrhea-inducing region of this viral enterotoxin. Therefore, developing an NSP4 lacking the enterotoxin domain could result in the introduction of a new subunit vaccine against rotaviruses in both humans and animals. Objectives: The aim of this study is the evaluation of rotavirus A NSP4 expression in E. coli expression system before and after removal of the diarrhea-inducing domain, which is the first step towards further immunological studies of the resulting protein. Materials and Methods: Splicing by overlap extension (SOEing) PCR was used to remove the diarrhea-inducing sequence from the NSP4 cDNA. Both the full-length (FL-NSP4) and the spliced (S-NSP4) cDNA amplicons were cloned into pET-32c and pGEX-6P-2. Expression levels of the recombinant proteins were evaluated in E. coli BL21 (DE3) by Western blot analysis. In addition, the toxicity of pET plasmids bearing the S-NSP4 and FL-NSP4 fragments was investigated by plasmid stability test. Results: For FL-NSP4, protein expression was detected for the strain containing the pGEX:FL-NSP4 plasmid, but not for the strain carrying pET:FL-NSP4. Hourly sampling up to 3 h showed that the protein production decreased by time. In contrast, expression of S-NSP4 was detected for pET:S-NSP4 strain, but not for pGEX:S-NSP4. Plasmid stability test showed that pET:S-NSP4 recombinant plasmid was almost stable, while pET:FL-NSP4 was unstable. Conclusions: This is the first report of production of rotavirus NSP4 lacking the diarrhea-inducing domain (S-NSP4). S-NSP4 shows less toxicity in this expression system and potentially could be a promising goal for rotavirus immunological and vaccine studies in the future.
https://www.ijbiotech.com/article_14131_65e0497ffe7341d2b06ce5bdb29b4acb.pdf
2016-06-01
50
57
10.15171/ijb.1233
Diarrhea
Enterotoxin
Expression
NSP4
Rotavirus
Splicing by overlap extension PCR
Mehdi
Sahmani
m.sahmani@gmail.com
1
Department of Clinical Biochemistry and Genetics, Cellular and Molecular Research Center, Qazvin University of Medical Sciences, Qazvin, Iran
AUTHOR
Siavash
Azari
siavashazari@hotmail.com
2
Department of Biotechnology, School of Paramedical Sciences, Qazvin University of Medical Sciences, Qazvin, Iran
AUTHOR
Majid
Tebianian
mtebianian@yahoo.com
3
Department of Biotechnology, Razi Vaccine and Serum Research Institute, Karaj, Iran
AUTHOR
Nematollah
Gheibi
gheibi_n@yahoo.com
4
Cellular and Molecular Research Center, Qazvin University of Medical Sciences, Qazvin, Iran
AUTHOR
Farzaneh
Pourasgari
farzaneh.pourasgari@gmail.com
5
Department of Biotechnology, Razi Vaccine and Serum Research Institute, Karaj, Iran
LEAD_AUTHOR
1. Rotavirus vaccines WHO position paper: January 2013- Recommendations. Vaccine. 2013;31(52):6170-6171. DOI: 10.1016/j.vaccine.2013.05.037
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7. Rajasekaran D, Sastri NP, Marathahalli JR, Indi SS, Pamidimukkala K, Suguna K, et al. The flexible C terminus of the rotavirus nonstructural protein NSP4 is an important determinant of its biological properties. J Gen Virol. 2008;89(6):1485-1496. DOI: 10.1099/vir.0.83617-0
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12. Zhang M, Zeng CQ, Morris AP, Estes MK. A functional NSP4 enterotoxin peptide secreted from rotavirus-infected cells. J Virol. 2000;74(24):11663-11670. DOI: 10.1128/JVI.74.24.11663-11670.2000
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13. Morris AP, Scott JK, Ball JM, Zeng CQ-Y, O’Neal WK, Estes MK. NSP4 elicits age-dependent diarrhea and Ca2+-mediated I- influx into intestinal crypts of CF mice. Am J Physiol. 1999;277(2):G431-G444
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14. Johansen K, Hinkula J, Espinoza F, Levi M, Zeng C, Ruden U, et al. Humoral and cell-mediated immune responses in humans to the NSP4 enterotoxin of rotavirus. J Med Virol. 1999;59(3):369-377. DOI: 10.1002/(SICI)1096-9071(199911)59:33.0.CO;2-N
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15. Browne EP, Bellamy AR, Taylor JA. Membrane-destabilizing activity of rotavirus NSP4 is mediated by a membrane-proximal amphipathic domain. J Gen Virol. 2000;81(8):1955-1959. DOI: 10.1099/0022-1317-81-8-1955
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16. Enouf V, Langella P, Commissaire J, Cohen J, Corthier G. Bovine rotavirus nonstructural protein 4 produced by Lactococcus lactis is antigenic and immunogenic. Appl Environ Microbiol. 2001;67(4):1423-1428. DOI: 10.1128/AEM.67.4.1423-1428.200 1
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17. Ray P, Malik J, Singh RK, Bhatnagar S, Bahl R, Kumar R, et al. Rotavirus nonstructural protein NSP4 induces heterotypic antibody responses during natural infection in children. J Infect Dis. 2003;187(11):1786-1793. DOI: 10.1086/375243
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18. Hyser JM, Zeng CQ, Beharry Z, Palzkill T, Estes MK. Epitope mapping and use of epitope-specific antisera to characterize the VP5* binding site in rotavirus SA11 NSP4. Virology. 2008;373(1):211-228. DOI: 10.1016/j.virol.2007.11.021
18
19. Deepa R, Durga Rao C, Suguna K. Structure of the extended diarrhea-inducing domain of rotavirus enterotoxigenic protein NSP4. Arch Virol. 2007;152(5):847-859. DOI: 10.1007/s00705-006-0921-x
19
20. Rajasekaran D, Sastri NP, Marathahalli JR, Indi SS, Pamidimukkala K, Suguna K, et al. The flexible C terminus of the rotavirus non-structural protein NSP4 is an important determinant of its biological properties. J Gen Virol. 2008;89(6):1485-1496. DOI: 10.1099/vir.0.83617-0
20
21. Vizzi E, Calvino E, Gonzalez R, Perez-Schael I, Ciarlet M, Kang G, et al. Evaluation of serum antibody responses against the rotavirus nonstructural protein NSP4 in children after rhesus rotavirus tetravalent vaccination or natural infection. Clin Diagn Lab Immunol. 2005;12(10):1157-1163. DOI: 10.1128/CDLI.12.10.1157-1163.2005
21
22. Hasenack BS, Botelho MVJ, Lauretti F, Melo FLd, Orlandi JM, Linhares REC, et al. The effect of concanavalin A on the replication of rotavirus (SA-11) in cell culture. Braz Arch Biol Technol. 2002;45:125-135. DOI: 10.1590/S1516-89132002000200003
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23. Pourasgari F, Ahmadian S, Hassan ZM, Mahdavi M, Salmanian AH, Sarbolouki MN, et al. Intranasal immunization of mice with VP2 DNA of human rotavirus a induces cellular and humoral immunity. Acta virol. 2008;52(4):225-229.
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24. Arnold M, Patton JT, McDonald SM. Culturing, storage and quantification of rotaviruses. Curr Protoc Microbiol. 2009; 15:C:15C.3:15C.3.1-15C.3.24. DOI: 10.1002/97804717292 59.mc1 5c03s15
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25. Pourasgari F, Ahmadian S, Salmanian AH. Expression and characterization of VP2 protein of human rotavirus a in a mammalian lung cell line. Acta Virol. 2007;51(4):261.
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29. Hyser JM, Collinson-Pautz MR, Utama B, Estes MK. Rotavirus disrupts calcium homeostasis by NSP4 viroporin activity. mBio. 2010;1(5):e00265-10. DOI: 10.1128/mBio.00265-10
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33. Pourasgari F, Ahmadian S, Salmanian AH, Sarbolouki MN, Massumi M. Low cytotoxicity effect of dendrosome as an efficient carrier for rotavirus VP2 gene transferring into a human lung cell line: dendrosome, as a novel intranasally gene porter. Mol Biol Rep. 2009;36(1):105-109. DOI: 10. 1007/s11033-007-9157-4
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34. Marelli B, Magni C. A simple expression system for Lactococcus lactis and Enterococcus faecalis. World J Microbiol Biotechnol. 2010;26(6):999-1007. DOI: 10.1007/ s11274-009-0262-5
34
35. Newton K, Meyer JC, Bellamy AR, Taylor JA. Rotavirus nonstructural glycoprotein NSP4 alters plasma membrane permeability in mammalian cells. J Virol. 1997;71(12):9458-9465.
35
ORIGINAL_ARTICLE
Phylogenetic Analysis of Aedes aegypti Based on Mitochondrial ND4 Gene Sequences in Almadinah, Saudi Arabia
Background: Aedes aegypti is the main vector of the yellow fever and dengue virus. This mosquito has become the major indirect cause of morbidity and mortality of the human worldwide. Dengue virus activity has been reported recently in the western areas of Saudi Arabia. There is no vaccine for dengue virus until now, and the control of the disease depends on the control of the vector. Objectives: The present study has aimed to perform phylogenetic analysis of Aedes aegypti based on mitochondrial NADH dehydrogenase subunit 4 (ND4) gene at Almadinah, Saudi Arabia in order to get further insight into the epidemiology and transmission of this vector. Materials and Methods: Mitochondrial ND4 gene was sequenced in the eight isolated Aedes aegypti mosquitoes from Almadinah, Saudi Arabia, sequences were aligned, and phylogenetic analysis were performed and compared with 54 sequences of Aedes reported in the previous studies from Mexico, Thailand, Brazil, and Africa. Results: Our results suggest that increased gene flow among Aedes aegypti populations occurs between Africa and Saudi Arabia. Conclusions: Phylogenetic relationship analysis showed two genetically distinct Aedes aegypti in Saudi Arabia derived from dual African ancestor.
https://www.ijbiotech.com/article_14134_c352342aadcaab1564b8d833431517dd.pdf
2016-06-01
58
62
10.15171/ijb.1329
Aedes
Mosquito
ND4 gene
Phylogenetic
Saudi Arabia
Khalil
AL ALI
alalikalil@yahoo.com
1
Department of Medical Laboratory Technology, College of Applied Medical Sciences, Taibah University, Almadinah Almanwra, Kingdom of Saudi Arabic
LEAD_AUTHOR
Ayman
El-Badry
aelbadry@kasralainy.edu.eg
2
Department of Medical Parasitology, Kasr Al-Ainy School of Medicine, Cairo University, Cairo, Egypt
AUTHOR
Mouhanad
AL ALI
alalimouhanad2013@gmail.com
3
Department of Institut Supérieur de la Santé et des Bioproduits d’Angers, Université d’Angers, Angers, France
AUTHOR
Wael
El-Sayed
waelsme@yahoo.com
4
Department of Microbiology, Faculty of Science, Ain Shams University, Cairo 11566, Egypt
Department of Biology, Faculty of Science, Taibah University, Almadinah Almunawarah 344, Saudi Arabia
AUTHOR
Hesham
El-Beshbishy
helbeshbishy@fakeeh.care
5
Department of Medical Laboratory Technology, College of Applied Medical Sciences, Taibah University, Almadinah Almanwra, Kingdom of Saudi Arabic
Department of Biochemistry, Faculty of Pharmacy, Al-Azhar University, Nasr City, Cairo, Egypt
AUTHOR
1. Forattini O. Culicidologia médica, vol. 2: Identificação, biologia, epidemiologia. São Paulo: Editora da Universidade de São Paulo. 2002
1
2. Taraphdar D, Sarkar A, Chatterjee S. Mass scale screening of common arboviral infections by an affordable, cost effective RT-PCR method. Asian Pac J Trop Biomed. 2012;2(2):97-101. DOI: 10.1016/S2221-1691(11)60200-1
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3. Brady OJ, Gething PW, Bhatt S, Messina JP, Brownstein JS, Hoen AG, et al. Refining the global spatial limits of dengue virus transmission by evidence-based consensus. PLoS Negl Trop Dis. 2012;6(8):e1760. DOI: 10.1371/journal.pntd.000 1760
3
4. El-Badry AA, El-Beshbishy HA, Al-Ali KH, Al-Hejin AM, El-Sayed WSM. Molecular and seroprevalence of imported dengue virus infection in Al-Madinah, Saudi Arabia. Comp Clin Path. 2013;23(4):861-868. DOI: 10.1007/s00580-013-1704-x
4
5. Azhar EI, Hashem AM, El-Kafrawy SA, Abol-Ela S, Abd-Alla AM, Sohrab SS, et al. Complete genome sequencing and phylogenetic analysis of dengue type 1 virus isolated from Jeddah, Saudi Arabia. Virol J. 2015;12:1. DOI: 10.1186/s 12985-014-0235-7
5
6. Bosio CF, Harrington LC, Jones JW, Sithiprasasna R, Norris DE, Scott TW. Genetic structure of Aedes aegypti populations in Thailand using mitochondrial DNA. Am J Trop Med Hyg. 2005;72(4):434-442.
6
7. Rattanarithikul R, Harrison BA, Panthusiri P, Coleman RE. Illustrated keys to the mosquitoes of Thailand I. Background; geographic distribution; lists of genera, subgenera, and species; and a key to the genera. Southeast Asian J Trop Med Public Health. 2005;36(1):1-80.
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8. Rattanarithikul R, Harbach RE, Harrison BA, Panthusiri P, Coleman RE, Richardson JH. Illustrated keys to the mosquitoes of Thailand. VI. Tribe Aedini. Southeast Asian J Trop Med Public Health. 2010;41(1):1-225.
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9. Jinbo U, Kato T, Ito M. Current progress in DNA barcoding and future implications for entomology. J Entomol Sci. 2011;14(2):107-124. DOI: 10.1111/j.1479-8298.2011.00449 .x
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10. Huber K, Le Loan L, Hoang TH, Ravel S, Rodhain F, Failloux AB. Genetic differentiation of the dengue vector, Aedes aegypti (Ho Chi Minh City, Vietnam) using microsatellite markers. Mol Ecol. 2002;11(9):1629-1635. DOI: 10.1046/j. 1365-294X.2002.01555.x
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11. Paupy C, Chantha N, Reynes JM, Failloux AB. Factors influencing the population structure of Aedes aegypti from the main cities in Cambodia. Heredity 2005;95(2):144-147. DOI: 10.1038/sj.hdy.6800698
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12. Scarpassa VM, Cardoza TB, Cardoso Junior RP. Population genetics and phylogeography of Aedes aegypti (Diptera: Culicidae) from Brazil. Am J Trop Med Hyg. 2008;78(6):895-903.
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13. Wesson DM, Porter CH, Collins FH. Sequence and secondary structure comparisons of ITS rDNA in mosquitoes (Diptera: Culicidae). Mol Phylogenet Evol. 1992;1(4):253-269. DOI: 10.1016/1055-7903(92)90001-W
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14. Marrelli MT, Sallum MA, Marinotti O. The second internal transcribed spacer of nuclear ribosomal DNA as a tool for Latin American anopheline taxonomy-a critical review. Mem Inst Oswaldo Cruz. 2006;101(8):817-832. DOI: http://dx.doi. org/10.1590/S0074-02762006000800002
14
15. Chan A, Chiang LP, Hapuarachchi HC, Tan CH, Pang SC, Lee R, et al. DNA barcoding: complementing morphological identification of mosquito species in Singapore. Parasit Vectors. 2014;7:569. DOI: 10.1186/s13071-014-0569-4
15
16. Urdaneta-Marquez L, Bosio C, Herrera F, Rubio-Palis Y, Salasek M, Black WCt. Genetic relationships among Aedes aegypti collections in Venezuela as determined by mitochondrial DNA variation and nuclear single nucleotide polymorphisms. Am J Trop Med Hyg. 2008;78(3):479-491.
16
17. Seixas G, Salgueiro P, Silva AC, Campos M, Spenassatto C, Reyes-Lugo M, et al. Aedes aegypti on Madeira Island (Portugal): genetic variation of a recently introduced dengue vector. Mem Inst Oswaldo Cruz. 2013;108(1):3-10. DOI: http: //dx.doi.org/10.1590/0074-0276130386
17
18. Da Costa-Da-Silva AL, Capurro ML, Bracco JE. Genetic lineages in the yellow fever mosquito Aedes (Stegomyia) aegypti (Diptera: Culicidae) from Peru. Mem Inst Oswaldo Cruz. 2005;100(6):539-544. DOI: http://dx.doi.org/10.1590/S0074-02762005000600007
18
19. Bracco JE, Capurro ML, Lourenco-de-Oliveira R, Sallum MA. Genetic variability of Aedes aegypti in the Americas using a mitochondrial gene: evidence of multiple introductions. Mem Inst Oswaldo Cruz. 2007;102(5):573-580. DOI: http://dx. doi.org/10.1590/S0074-02762007005000062
19
20. Paduan Kdos S, Ribolla PE. Mitochondrial DNA polymorphism and heteroplasmy in populations of Aedes aegypti in Brazil. J Med Entomol. 2008;45(1):59-67.
20
21. Schaffner F. Les moustiques d’Europe logiciel d’identification et d’enseignement; an identification and training programme; francais, english = The mosquitoes of Europe. Paris: IRD Éditions; 2001
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22. Collins FH, Mendez MA, Rasmussen MO, Mehaffey PC, Besansky NJ, Finnerty V. A ribosomal RNA gene probe differentiates member species of the Anopheles gambiae complex. Am J Trop Med Hyg. 1987;37(1):37-41.
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23. Kumar S, Tamura K, Nei M. MEGA3: Integrated software for Molecular Evolutionary Genetics Analysis and sequence alignment. Brief Bioinform. 2004;5(2):150-163. DOI: 10. 1093/bib/5.2.150
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24. Paupy C, Le Goff G, Brengues C, Guerra M, Revollo J, Barja Simon Z, et al. Genetic structure and phylogeography of Aedes aegypti, the dengue and yellow-fever mosquito vector in Bolivia. Infection, genetics and evolution. Infect Genet Evol. 2012;12(6):1260-1269. DOI: 10.1016/j.meegid.2012.04.012
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25. Moore M, Sylla M, Goss L, Burugu MW, Sang R, Kamau LW, et al. Dual African origins of global Aedes aegypti s.l. populations revealed by mitochondrial DNA. PLoS Negl Trop Dis. 2013;7(4):e2175. DOI: 10.1371/journal.pntd.0002175
25
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26
ORIGINAL_ARTICLE
Cloning, Codon Optimization, and Expression of Yersinia intermedia Phytase Gene in E. coli
Background: Phytate is an anti-nutritional factor in plants, which catches the most phosphorus contents and some vital minerals. Therefore, Phytase is added mainly as an additive to the monogastric animals’ foods to hydrolyze phytate and increase absorption of phosphorus. Objectives: Y. intermedia phytase is a new phytase with special characteristics such as high specific activity, pH stability, and thermostability. Our aim was to clone, express, and characterizea codon optimized Y. intermedia phytase gene in E. coli. Materials and Methods: The Y. intermedia phytase gene was optimized according to the codon usage in E. coli. The sequence was synthesized and sub-cloned in pET-22b (+) vector and transformed into E. coli Bl21 (DE3). The protein was expressed in the presence of IPTG at a final concentration of 1 mM at 30°C. The purification of recombinant protein was performed by Ni2+ affinity chromatography. Phytase activity and stability were determined in various pH and temperatures. Results: The codon optimized Y. intermedia phytase gene was sub-cloned successfully.The expression was confirmed by SDS-PAGE and Western blot analysis. The recombinant enzyme (approximately 45 kDa) was purified. Specific activity of enzyme was 3849 (U.mg-1) with optimal pH 5 and optimal temperature of 55°C. Thermostability (80°C for 15 min) and pH stability (3-6) of the enzyme were 56 and more than 80%, respectively. Conclusions: The results of the expression and enzyme characterization revealed that the optimized Y. intermedia phytase gene has a good potential to be produced commercially andto be applied in animals’ foodsindustry.
https://www.ijbiotech.com/article_14136_ef6f9f044a1b735f7804bc45da7d693f.pdf
2016-06-01
63
69
10.15171/ijb.1412
Codon optimization
Expression in E. coli
Rare codons
Y. intermedia phytase
Maryam
Mirzaei
mirzaei18@gmail.com
1
Department of Biology, Master of Science, Faculty of Science, Shahrekord University, Shahrekord, Iran
AUTHOR
Behnaz
Saffar
biotechresearch2014@gmail.com
2
Department of Genetics, Faculty of Sciences, Shahrekord University, Shahrekord, Iran
LEAD_AUTHOR
Behzad
Shareghi
b_shareghi@yahoo.com
3
Department of Biology, Faculty of Science, Shahrekord University, Shahrekord, Iran
AUTHOR
1. Sapna JJ, Singh B. Characteristics and biotechnological applications of bacterial phytases. Process Biochem. 2016;51: 159-169. DOI: 10.1016/j.procbio.2015.12.004
1
2. Corrêa TLR, Vieira de Queiroz M, Fernandes de Araújo E. Cloning, recombinant expression and characterization of a new phytase from Penicillium chrysogenum. Microbiol Res. 2015;170:205-212. DOI: 10.1016/j.micres.2014.06.005
2
3. García-Mantrana I, Yebra MJ, Haros M, Monedero V. Expression of bifidobacterial phytases in Lactobacillus casei and their application in a food model of whole-grain sourdough bread. Int. J Food Microb. 2016:4:18-24. DOI: 10.1016/j.ijfoodmicro.2015.09.003
3
4. Tamim N, Angel R, Christman M. Influence of dietary calcium nd phytase on phytate phosphorus hydrolysis in broiler chickens. Poult Sci. 2004;83(8):1358-1367. DOI:10.1093/ps/83.8.1358
4
5. Schlemmer U, Frølich W, Prieto RM, Grases F. Phytate in foods and significance for humans: food sources, intake, processing, bioavailability, protective role and analysis. Mol Nutr Food Res. 2009;53(2):330-375. DOI: 10.1002/mnfr.200900099
5
6. Fuhrman J, Zhang H, Schroder J, Davis R, Payton M. Water-soluble phosphorus as affected by soil to extractant Ratios, extraction times, and electrolyte. Commun Soil Sci Plant Anal. 2005,36(7):925-935. DOI: 10.1081/CSS-200049482
6
7. Vasudevan UM, Salim SHB, Pandey A. A comparative analysis of recombinant expression and solubility screening of two phytases in E. coli. Food Technol Biotech. 2011;49(3):304-309.
7
8. Yao MZ, Zhang YH, Lu WL, Hu MQ, Wang W, Liang AH. Phytases: crystal structures, protein engineering and potential biotechnological applications. J Appl Microbiol. 2012;112(1):1-14.
8
9. Sajidan A, Farouk A, Greiner R, Jungblut P, Müller EC, Borriss R. Molecular and physiological characterisation of a 3-phytase from soil bacterium Klebsiella sp. ASR1. Appl Microbiol Biotechnol. 2004;65(1):110-118. DOI: 10.1007/s 00253-003-1530-1
9
10. Lei XG, Stahl CH. Biotechnological development of effective phytases for mineral nutrition and environmental protection. Appl Microbiol Biotechnol. 2001;57(4):474-481. DOI: 10.1007/s002530100795
10
11. Luo H, Yao B, Yuan T, Wang Y, Shi X, Wu N, et al. Overexpression of Escherchia coli phytase with high specific activity. Sheng Wu Gong Cheng Xue Bao. 2004;20:78-84.
11
12. Puigbò P, Guzmán E, Romeu A, Garcia-Vallvé S. OPTIMIZER: a web server for optimizing the codon usage of DNA sequences. Nuc Acids Res. 2007;35:126-131. DOI: 10.1093/nar/gkm219
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13. Li G, Zhu J, Sun J, Wu Z, Chen J, Yan J, et al. Cloning of the phytase gene phyA from Aspergillus ficuum 3.4322 and its expression in yeast. Fungal Divers. 2003;13:85-93
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14. Kim YO, Lee JK, Kim HK, Yu JH, Oh TK. Cloning of the thermostable phytase gene (phy) from Bacillus sp. DS11 and its overexpression in E. coli. FEMS Microbiol Lett. 1998;162(1):185-191. DOI: 10.1111/j.1574-6968.1998.tb 12997.x
14
15. Huang H, Luo H, Yang, P, Meng, K, Wang, Y, Yuan, T, et al. A novel phytase with preferable characteristics from Yersinia intermedia. Biochem Biophys Res Commun. 2006;350(4): 884-889. DOI: 10.1016/j.bbrc.2006.09.118
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16. Sambrook JRD. “Molecular cloning; a laboratory manual. Cold Spring Harbor Laboratory Press, Cold Spring Harbor.” New York. 2001. DOI: 10.1086/428170
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