Expression and Purification of a Novel Computationally Designed Antigen for Simultaneously Detection of HTLV-1 and HBV Antibodies
Abstract
Computational tools are reliable alternatives to laborious work in chimeric protein design. In this study, a chimeric antigen was designed using computational techniques for simultaneous detection of anti-HTLV-I and anti-HBV in infected sera. Databases were searched for amino acid sequences of HBV/HLV-I diagnostic antigens. The immunodominant fragments were selected based on propensity scales. The diagnostic antigen was designed using these fragments. Secondary and tertiary structures were predicted and the B-cell epitopes were mapped on the surface of built model. The synthetic DNA coding antigen was sub-cloned into pGS21a expression vector. SDS-PAGE analysis showed that glutathione fused antigen was highly expressed in E. coli BL21 (DE3) cells. The recombinant antigen was purified by nickel affinity chromatography. ELISA results showed that soluble antigen could specifically react with the HTLV-I and HBV infected sera. This specific antigen could be used as suitable agent for antibody-antigen based screening tests and can help clinicians in order to perform quick and precise screening of the HBV and HTLV-I infections.
1. Laperche S. Multinational assessment of blood‐borne virus testing and transfusion safety on the African continent. Transfusion 2013; 53(4):816-26.
2. Mehta SH, Astemborski J, Kirk GD, Strathdee SA, Nelson KE, Vlahov D, et al. Changes in blood-borne infection risk among injection drug users. J Infect Dis 2011; 203(5):587-94.
3. Rafatpanah H, Hedayati-Moghaddam MR, Fathimoghadam F, Bidkhori HR, Shamsian SK, Ahmadi S, et al. High prevalence of HTLV-I infection in Mashhad, Northeast Iran: a population- based seroepidemiology survey. J Clin Virol 2011;52(3):172-6.
4. Azarpazhooh MR, Hasanpour K, Ghanbari M, Rezaee SR, Mashkani B, Hedayati-Moghaddam MR, et al. Human T-lymphotropic virus type 1 prevalence in Northeastern Iran, Sabzevar: an epidemiologic-based study and phylogenetic analysis. AIDS Res Hum Retroviruses 2012; 28(9):1095-101.
5. Fathimoghaddam F, Hedayati-Moghaddam MR, Bidkhori HR, Ahmadi S, Sima HR. The prevalence of hepatitis B antigen-positivity in the general population of Mashhad, Iran. Hepat Mon 2011; 11(5):346-50.
6. Shakeri MT, Foghanian B, Nomani H, Ghayour- Mobarhan M, Nabavinia MS, Rostami S, et al. The Prevalence of Hepatitis B Virus Infection in Mashhad, Iran: A Population-Based Study. Iran Red Crescent Med J 2013; 15(3):245-8.
7. Allain JP. Occult hepatitis B virus infection: implications in transfusion. Vox sanguinis 2004; 86(2):83-91.
8. Proietti FA, Carneiro-Proietti ABF, Catalan-Soares BC, Murphy EL. Global epidemiology of HTLV-I infection and associated diseases. Oncogene 2005; 24(39):6058-68.
9. Gallo D, Penning LM, Hanson C. Detection and differentiation of antibodies to human T-cell lymphotropic virus types I and II by the immunofluorescence method. J Clin Microbiol 1991;
29(10):2345-7.
10. Wang JT, Lee CZ, Chen PJ, Wang TH, Chen DS.Transfusion‐transmitted HBV infection in an endemic area: the necessity of more sensitive screening for HBV carriers. Transfusion 2002; 42(12):1592-7.
11. . Caterino-de-Araujo A, de los Santos-Fortuna E, Meleiro MC, Suleiman J, Calabro ML, Favero A, et al. Sensitivity of two enzyme-linked immunosorbent assay tests in relation to western blot in detecting human T-cell lymphotropic virus types I and II infection among HIV-1 infected patients from Sao Paulo, Brazil. Diagn Microbiol Infect Dis. 1998;30(3):173-82.
12. Diepolder HM. Clearance of HBV markers and HCC risk: who is safe? Gut 2014.
13. Rahbar MR, Rasooli I, Gargari SLM, Sandstrom G, Amani J, Fattahian Y, et al. A potential in silico antibody–antigen based diagnostic test for precise identification of Acinetobacter baumannii. J Theor Biol 2012; 294:29-39.
14. Edgar RC. MUSCLE: multiple sequence alignment with high accuracy and high throughput. Nucleic acids research 2004; 32(5):1792-7.
15. Gouet P, Robert X, Courcelle E. ESPript/ENDscript: extracting and rendering sequence and 3D information from atomic structures of proteins. Nucleic Acids Res 2003; 31(13):3320-3.
16. Roy A, Kucukural A, Zhang Y. I-TASSER: a unified platform for automated protein structure and function prediction. Nat Protoc 2010; 5(4):725-38.
17. Wiederstein M, Sippl MJ. ProSA-web: interactive web service for the recognition of errors in three-dimensional structures of proteins. Nucleic Acids Res 2007; 35(suppl 2):W407-10.
18. Larsen JE, Lund O, Nielsen M. Improved method for predicting linear B-cell epitopes. Immunome Res 2006;2:2.
19. Doytchinova IA, Flower DR. VaxiJen: a server for prediction of protective antigens, tumour antigens and subunit vaccines. BMC Bioinformatics 2007; 8:4.
20. Davis GD, Elisee C, Newham DM, Harrison RG. New fusion protein systems designed to give soluble expression in Escherichia coli. Biotechnol Bioeng 1999;65(4):382-8.
21. Smedile A, Lavarini C, Crivelli O, Raimondo G, Fassone M, Rizzetto M. Radioimmunoassay detection of igm antibodies to the hbv‐associated delta (δ) antigen: Clinical significance in δ infection. J Med Virol 1982; 9(2):131-8.
22. Dipti CA, Jain S, Navin K. A novel recombinant multiepitope protein as a hepatitis C diagnostic intermediate of high sensitivity and specificity. Protein Expr Purif 2006; 47(1):319-28.
23. Thorstensson R, Albert J, Andersson S. Strategies for diagnosis of HTLV‐I and‐II. Transfusion 2002;42(6):780-91.
24. Zhao L, Rai SK, Grosmaire LS, Ledbetter JA, Fell HP.Construction, expression, and characterization of anticarcinoma sFv fused to IL-2 or GM-CSF. J Hematother Stem Cell Res 1999; 8(4):393-9.
25. Zhang Y, Song S, Liu C, Wang Y, Xian X, He Y, et al.Generation of chimeric HBc proteins with epitopes in E. coli: Formation of virus-like particles and a potent inducer of antigen-specific cytotoxic immune response and anti-tumor effect in vivo. Cell Immunol 2007;247(1):18-27.
26. Lillehoj E, Alexander S, Dubrule C, Wiktor S, Adams R,Tai C-C, et al. Development and evaluation of a human T-cell leukemia virus type I serologic confirmatory assay incorporating a recombinant envelope polypeptide. J Clin Microbiol 1990; 28(12):2653-8.
27. Kopito RR. Aggresomes, inclusion bodies and protein aggregation. Trends Cell Biol 2000; 10(12):524-30.
28. Sharma SK, Suresh MR, Wuest FR. Improved soluble expression of a single-chain antibody fragment in E. coli for targeting CA125 in epithelial ovarian cancer. Protein Expr Purif. 2014; 29:27-37.
29. Tolia NH, Joshua-Tor L. Strategies for protein coexpression in Escherichia coli. Nat Methods 2006;3(1):55-64.
30. Frikha-Gargouri O, Gdoura R, Znazen A, Gargouri B, Gargouri J, Rebai A, et al. Evaluation of an in silico predicted specific and immunogenic antigen from the OmcB protein for the serodiagnosis of Chlamydia trachomatis infections. BMC Microbiol 2008;8(1):217.Saha S, Raghava G. Prediction of continuous B‐cell epitopes in an antigen using recurrent neural network. Proteins 2006; 65(1):40-8.
| Files | ||
| Issue | Vol 14, No 2 (2015) | |
| Section | Articles | |
| Keywords | ||
| Blood screening Computational analysis HBV HTLV-I Transfusion | ||
| Rights and permissions | |
|
This work is licensed under a Creative Commons Attribution-NonCommercial 4.0 International License. |
