Volume 25, Issue 11 (2-2019)                   RJMS 2019, 25(11): 52-60 | Back to browse issues page

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abdollahi M, Honari H, e'temad aubi S M. Expression of SO6-STxB gene cassette in Escherichia coli and investigation of antibody titer. RJMS 2019; 25 (11) :52-60
URL: http://rjms.iums.ac.ir/article-1-5164-en.html
Imam Hosein University, Tehran Iran , honari.hosein@gmail.com
Abstract:   (3193 Views)
Background: One of the ways to strengthen the effect of vaccines is the use of adjuvant. STxB has a carrier role and can act as an adjuvant; thus it can be fused with vaccine candidate antigens in order to produce efficient vaccines. Saponaria officinalis is a plant that has shown N-glycosidase activity. SO6 isoform of this plant, depurinates the adenine 4324 in the conserved sequence GAGA in 28SrRNA and disrupts protein synthesis. The aim of this study was expression of SO6-STxB gene in Escherichia coli (E. coli) and investigation of antibody titer in mice.
Methods: In this study SO6 gene with BamHI and SalI restriction enzyme sites were isolated from pUC57 plasmid and subcloned into pET28a (+) -STxB expression vector and transferred to E. coli BL21 (DE3). Expression of SO6-STxB gene cassette was induced by IPTG and purified by nickel affinity chromatography. The recombinant protein was confirmed by western blotting. Mice were immunized intraperitoneally with purified protein and serum IgG titers were measured by ELISA.
Results: Subcloning of SO6–STxB gene in pET28a (+) expression vector was confirmed by PCR and enzyme digestion reaction. A 37/5 kDa recombinant protein was confirmed by SDS-PAGE and western blotting. The antibody generated from mouse serum was isolated and confirmed by ELISA.
Conclusion: Purified recombinant antigen STxB-SO6 can be used for research and be a suitable anti-cancer vaccine candidates.
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Type of Study: Research | Subject: Biology

1. 1. Stirpe F, Battelli M. Ribosome-inactivating proteins: progress and problems. Cell Mol Life Sci; 2006. 63(16):1850-66.
2. 2. Ferreras J, Barbieri L, Girbés T, Battelli MG, Rojo MA, Arias FJ, et al. Distribution and properties of major ribosome-inactivating proteins (28 S rRNA N-glycosidases) of the plant Saponaria officinalis L.(Caryophyllaceae). Biochim Biophys Acta; 1993. 1216(1):31-42.
3. 3. Maras B, Ippoliti R, De Luca E, Lendaro E, Bellelli A, Barra D, et al. The amino acid sequence of a ribosome-inactivating protein from Saponaria officinalis seeds. Biochem Int; 1990. 21(5):831-8.
4. 4. Duggar BM, Armstrong JK. The effect of treating the virus of tobacco mosaic with the juices of various plants. Ann Missouri Bot Gard; 1925. 12(4):359-66.
5. 5. Leah R, Tommerup H, Svendsen I, Mundy J. Biochemical and molecular characterization of three barley seed proteins with antifungal properties. J Biol Chem; 1991. 266(3):1564-73.
6. 6. Wang S, Li Z, Li S, Di R, Ho CT, Yang G. Ribosome-inactivating proteins (RIPs) and their important health promoting property. RSC Adv; 2016. 6(52):46794-805.
7. 7. Bouter A, Delord B, Dransart E, Poirier C, Johannes L, Effenterre D. Intracellular trafficking of Shiga‐toxin‐B‐subunit‐functionalized spherulites. J Cell Biol; 2008. 100(12):717-28.
8. 8. Pina DG, Johannes L. Cholera and Shiga toxin B-subunits: thermodynamic and structural considerations for function and biomedical applications. Toxicon; 2005. 45(4):389-93.
9. 9. Janssen KP, Vignjevic D, Boisgard R, Falguières T, Bousquet G, Decaudin D, et al. In vivo tumor targeting using a novel intestinal pathogen-based delivery approach. J Cancer Res; 2006. 66(14):7230-6.
10. 10. Choi NW, Estes MK, Langridge WH, Oral immunization with a shiga toxin B subunit: rotavirus NSP4 90 fusion protein protects mice against gastroenteritis. Vaccine 2005;23(44): 5168-5.
11. 11-Marcato P, Thomas P, Griener, George L, Mulvey, Glen D. Armstrong2. Recombinant Shiga toxin B-subunit-keyhole limpet hemocyanin conjugate vaccine protects mice from Shigatoxemia. Infect Immun; 2005. 73(10):6523-9.
12. 12. Honari H, Amlashi I, Minaee ME, Safaee S. [Immunogenicity in guinea pigs by IpaD-STxB
13. recombinant protein]. AMUJ; 2013. 16(4):83-93. [Persian].
14. 13. Honari H, Amlashi I, Minaei ME. [Expression of recombinant proteins IpaD-STxB and immunogenicity STxB in the mice]. J Mazandaran Univ Med Sci; 2014. 23:196-206. [Persian].
15. 14. Honari H, Minaei HH, Ebrahim M. [Analyzing the various fusions for ctxB, ipaD and stxB genes of Shigella dysenteriae and Vibrio cholera by bioinformatics tools]. Genetics in the 3rd Millennium; 2013. 11(2):3070-7. [Persian].
16. 15. Abdollahi M, Honari H, Nazarian S, Masoudi Kerahroudi M. [Subcloning and expression of SO6 gene, Saponaria officinalis plant in E. coli and investigation of antibody titer in rats]. JSSU; 2017. 24(12):1024-33. [Persian].
17. 16. Sambrook J, Russell DW. Molecular cloning: a laboratory manual 3rd edition. Coldspring-Harbour Laboratory Press, UK. 2001.
18. 17. Tonello F, Pellizzari R, Pasqualato S, Grandi G, Peggion E, Montecucco C. Recombinant and truncated tetanus neurotoxin light chain: cloning, expression, purification, and proteolytic activity. Protein Expr. Purif; 1999. 15(2):221-7.
19. 18. Chia MY, Hsiao SH, Chan HT, Do YY, Huang PL, Chang HW, et al. The immunogenicity of DNA constructs co-expressing GP5 and M proteins of porcine reproductive and respiratory syndrome virus conjugated by GPGP linker in pigs. Vet Microbiol; 2010 Dec 15. 146(3):189-99.
20. 19. Honari H, Amlashi I, Minaei ME, Safaei S. [Immunogenicity in guinea pigs by IpaD-STxB recombinant protein]. AMUJ; 2013. 83-93. Persian.
21. 20. Honari H, Amlashi ????, Minaei ME. [Expression of recombinant proteins IpaD-STxB and immunogenicity STxB in the mice]. J Mazandaran Univ Med Sci; 2014. 183-93. [Persian].
22. 21. Baranvand M, Honari H. [Nasal immunogenicity induced by STxB and STxB-IpaD antigens in laboratory rats]. Koomesh; 2015. 397-403. [Persian].
23. 22. Abdollahi M, Honari H, Nazarian SH, Masoudi Kerahroudi M. [Subcloning and expression of SO6 gene, Saponaria officinalis plant in E. coli and investigation of antibody titer in laboratory rat]. J Shahid Sadoughi Univ Med Sci; 2017. 24(12):1024-33. [Persian].
24. 23. Walsh MJ, Dodd JE, Hautbergue GM. Ribosome-inactivating proteins: Potent poisons and molecular tools. Virulence; 2013. 4(8):774-84.
25. 24. Schrot J, Weng A, Melzig MF. Ribosome-inactivating and related proteins. Toxins; 2015. 7(5):1556-615.
26. 25. Bagga S, Hosur M, Batra JK. Cytotoxicity of ribosome‐inactivating protein saporin is not mediated through α2‐macroglobulin receptor. FEBS Lett; 2003. 541(1-3):16-20.
27. 26. Bergamaschi G, Perfetti V, Tonon L, Novella A, Lucotti C, Danova M, et al. Saporin, a ribosome‐inactivating protein used to prepare immunotoxins, induces cell death via apoptosis. Br J Haematol; 1996. 93(4):789-94.
28. 27. Bolognesi A, Tazzari PL, Olivieri F, Polito L, Falini B, Stirpe F. Induction of apoptosis by ribosome‐inactivating proteins and related immunotoxins. Int J Cancer; 1996. 68(3):349-55.
29. 28. Mir LM, Banoun H, Paoletti C. Introduction of definite amounts of nonpermeant molecules into living cells after electropermeabilization: direct access to the cytosol. Exp Cell Res; 1988. 175(1):15-25.
30. 29. Kodama T, Doukas AG, Hamblin MR. Delivery of ribosome-inactivating protein toxin into cancer cells with shock waves. Cancer Lett; 2003. 189(1):69-75.
31. 30. Selbo P, Kristian al, Høgset A, Prasmickaite L, Berg K. Photochemical internalisation: a novel drug delivery system. Tumor Biol; 2002. 23(2):103-12.
32. 31. Stirpe F. Ribosome-inactivating proteins. Toxicon; 2004. 44(4):371-83.

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