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Design and expression of polytopic construct of cathepsin-L1, SAP-2 and FhTP16.5 proteins of Fasciola hepatica

Published online by Cambridge University Press:  04 March 2020

S. Aghamolaei ,
B. Kazemi ,
M. Bandehpour ,
M.M. Ranjbar ,
S. Rouhani ,
A. Javadi Mamaghani  and
S.J.S. Tabaei Open the ORCID record for S.J.S. Tabaei [Opens in a new window]
S. Aghamolaei
Affiliation:
Department of Parasitology and Mycology, School of Medicine, Student Research Committee, Shahid Beheshti University of Medical Sciences, Tehran, Iran
B. Kazemi
Affiliation:
Cellular & Molecular Biology Research Center, School of Medicine, Shahid Beheshti University of Medical Sciences, Tehran, Iran
M. Bandehpour
Affiliation:
Cellular & Molecular Biology Research Center, School of Medicine, Shahid Beheshti University of Medical Sciences, Tehran, Iran
M.M. Ranjbar
Affiliation:
Razi Vaccine and Serum Research Institute, Agricultural Research, Education and Extension Organization (AREEO), Karaj, Iran
S. Rouhani
Affiliation:
Department of Parasitology and Mycology, School of Medicine, Shahid Beheshti University of Medical Sciences, Tehran, Iran
A. Javadi Mamaghani
Affiliation:
Department of Parasitology and Mycology, School of Medicine, Student Research Committee, Shahid Beheshti University of Medical Sciences, Tehran, Iran
S.J.S. Tabaei*
Affiliation:
Department of Parasitology and Mycology, School of Medicine, Shahid Beheshti University of Medical Sciences, Tehran, Iran
*
Author for correspondence: S.J.S. Tabaei, E-mail: seyyedtabaei@gmail.com
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Abstract

The enzyme-linked immunosorbent assay (ELISA) technique can play an important role in the early detection of fascioliasis. However, they have some diagnostic limitations, including cross-reaction with other helminths. It seems that the combination of recombinant parasite proteins as antigen can reduce these problems. Hence, the present study was aimed to design and confirm the antigenic recombinant multi-epitope (rMEP) construct of three protein epitopes (linear and conformational B-cell epitopes) of the parasite using immunoinformatic tools. For this purpose, the tertiary structures of Fasciola hepatica cathepsin-L1, saposin-like protein 2 and 16.5-kDa tegument-associated protein were predicted using the I-TASSER server. Validation of the modelled structures was performed by Ramachandran plots. The antigenic epitopes of the proteins were achieved by analysing the features of the IEDB server. The synthesized gene was cloned into the pET-22b (+) expression vector and transformed into the Escherichia coli BL21. Sodium dodecyl sulfate polyacrylamide gel electrophoresis was used to verify and analyse the expression of the rMEP protein. Western blotting was utilized to confirm rMEP protein immunogenicity in two forms, one using an anti-His tag antibody and the other with human pooled sera samples (fascioliasis, non-fascioliasis and negative control sera). Our results demonstrated that the rMEP designed for the three proteins of F. hepatica was highly antigenic, and immune-detection techniques confirmed the antigen specificity. In conclusion, the presented antigenic multi-epitope may be very helpful to develop serodiagnostic kits such as indirect ELISA to evaluate the proper diagnosis of fascioliasis in humans and ruminants.

Keywords

FascioliasisFasciolaepitopescathepsin-L1SAP-2FhTP16.5recombinant proteinsmulti-epitopeimmunoinformatic
Type
Research Paper
Information
Journal of Helminthology , Volume 94 , 2020 , e134
Copyright
Copyright © Cambridge University Press 2020

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References

Abdalrahman, RA, Ahmed, SS, Elnaeem, MA, et al. (2019) Translationally controlled tumor protein TCTP as peptide vaccine against Schistosoma japonicum: an immunoinformatics approach. bioRxiv, 466847.Google Scholar
Adela Valero, M, Victoria Periago, M, Pérez-Crespo, I, Rodríguez, E, Jesús Perteguer, M, Gárate, T, González-Barberá, EM and Mas-Coma, S (2012) Assessing the validity of an ELISA test for the serological diagnosis of human fascioliasis in different epidemiological situations. Tropical Medicine & International Health 17, 630–636.CrossRefGoogle Scholar
Adhikari, UK, Tayebi, M and Rahman, MM (2018) Immunoinformatics approach for epitope-based peptide vaccine design and active site prediction against polyprotein of emerging oropouche virus. Journal of Immunology Research 2018, 6718083.CrossRefGoogle ScholarPubMed
Alavi, SM and Salmanzadeh, S (2016) Schistosomiasis in Iran, From the past till elimination. International Journal of Infection 3, e36075.CrossRefGoogle Scholar
Ali, M, Pandey, RK, Khatoon, N, Narula, A, Mishra, A and Prajapati, VK (2017) Exploring dengue genome to construct a multi-epitope based subunit vaccine by utilizing immunoinformatics approach to battle against dengue infection. Scientific Reports 7, 9232.CrossRefGoogle ScholarPubMed
Amer, S, ElKhatam, A, Zidan, S, Feng, Y and Xiao, L (2016) Identity of Fasciola spp. in sheep in Egypt. Parasites & Vectors 9, 623.CrossRefGoogle ScholarPubMed
Ansari, HR and Raghava, GP (2010) Identification of conformational B-cell epitopes in an antigen from its primary sequence. Immunome Research 6, 6.CrossRefGoogle Scholar
Assis, LMd, Sousa, J, Pinto, N, Silva, A, Vaz, AFdM, Andrade, PP, Carvalho, EMd and De Melo, M (2014) B-cell epitopes of antigenic proteins in Leishmania infantum: an in silico analysis. Parasite Immunology 36, 313–323.CrossRefGoogle ScholarPubMed
Bradford, MM (1976) A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Analytical Biochemistry 72, 248–254.CrossRefGoogle ScholarPubMed
Cabán-Hernández, K and Espino, AM (2013) Differential expression and localization of saposin-like protein 2 of Fasciola hepatica. Acta Tropica 128, 591–597.CrossRefGoogle ScholarPubMed
Cabán-Hernández, K, Gaudier, JF, Ruiz-Jiménez, C and Espino, AM (2014) Development of two antibody detection enzyme-linked immunosorbent assays for serodiagnosis of human chronic fascioliasis. Journal of Clinical Microbiology 52, 766–772.CrossRefGoogle ScholarPubMed
Chou, P and Fasman, GD (2009) Amino acid sequence. Advances in Enzymology and Related Areas of Molecular Biology 47, 45.Google Scholar
Cornelissen, JB, Gaasenbeek, CP, Boersma, W, Borgsteede, FH and van Milligen, FJ (1999) Use of a pre-selected epitope of cathepsin-L1 in a highly specific peptide-based immunoassay for the diagnosis of Fasciola hepatica infections in cattle. International Journal for Parasitology 29, 685–696.CrossRefGoogle Scholar
Cornelissen, JB, Gaasenbeek, CP, Borgsteede, FH, Holland, WG, Harmsen, MM and Boersma, WJ (2001) Early immunodiagnosis of fasciolosis in ruminants using recombinant Fasciola hepatica cathepsin L-like protease. International Journal for Parasitology 31, 728–737.CrossRefGoogle ScholarPubMed
Cwiklinski, K, Allen, K, LaCourse, J, Williams, DJ, Paterson, S and Hodgkinson, JE (2015) Characterisation of a novel panel of polymorphic microsatellite loci for the liver fluke, Fasciola hepatica, using a next generation sequencing approach. Infection, Genetics and Evolution 32, 298–304.CrossRefGoogle ScholarPubMed
Emini, EA, Hughes, JV, Perlow, D and Boger, J (1985) Induction of hepatitis A virus-neutralizing antibody by a virus-specific synthetic peptide. Journal of Virology 55, 836–839.CrossRefGoogle ScholarPubMed
Espino, AM and Hillyer, GV (2003) Molecular cloning of a member of the Fasciola hepatica saposin-like protein family. Journal of Parasitology 89, 545–552.CrossRefGoogle ScholarPubMed
Espinoza, J, Timoteo, O and Herrera-Velit, P (2005) Fas2-ELISA in the detection of human infection by Fasciola hepatica. Journal of Helminthology 79, 235–240.CrossRefGoogle ScholarPubMed
Ezzat, RF, Karboli, TA, Kasnazani, KA and Hamawandi, AM (2010) Endoscopic management of biliary fascioliasis: a case report. Journal of Medical Case Reports 4, 83.CrossRefGoogle ScholarPubMed
Figueroa-Santiago, O, Delgado, B and Espino, AM (2011) Fasciola hepatica saposin-like protein-2–based ELISA for the serodiagnosis of chronic human fascioliasis. Diagnostic Microbiology and Infectious Disease 70, 355–361.CrossRefGoogle ScholarPubMed
Gaudier, JF, Cabán-Hernández, K, Osuna, A and Espino, AM (2012) Biochemical characterization and differential expression of a 16.5-kilodalton tegument-associated antigen from the liver fluke Fasciola hepatica. Clinical and Vaccine Immunology 19, 325–333.CrossRefGoogle ScholarPubMed
Gottstein, B, Schneeberger, M, Boubaker, G, Merkle, B, Huber, C, Spiliotis, M, Müller, N, Garate, T and Doherr, MG (2014) Comparative assessment of ELISAs using recombinant saposin-like protein 2 and recombinant cathepsin L-1 from Fasciola hepatica for the serodiagnosis of human fasciolosis. PLoS Neglected Tropical Diseases 8, e2860.CrossRefGoogle ScholarPubMed
Ikeda, T, Oikawa, Y and Nishiyama, T (1996) Enzyme-linked immunosorbent assay using cysteine proteinase antigens for immunodiagnosis of human paragonimiasis. The American Journal of Tropical Medicine and Hygiene 55, 434–437.CrossRefGoogle ScholarPubMed
Intapan, PM, Tantrawatpan, C, Maleewong, W, Wongkham, S, Wongkham, C and Nakashima, K (2005) Potent epitopes derived from Fasciola gigantica cathepsin L1 in peptide-based immunoassay for the serodiagnosis of human fascioliasis. Diagnostic Microbiology and Infectious Disease 53, 125–129.CrossRefGoogle ScholarPubMed
Jaros, S, Zygner, W, Januszkiewicz, K, Jaros, D and Wędrychowicz, H (2010) Molecular cloning and bioinformatic characterisation of FhPcW1, a new isoform of cathepsin L from adult Fasciola hepatica. Bulletin of the Veterinary Institute in Pulawy 54, 585–590.Google Scholar
Karplus, P and Schulz, G (1985) Prediction of chain flexibility in proteins. Naturwissenschaften 72, 212–213.CrossRefGoogle Scholar
Koç, Z, Ulusan, S and Tokmak, N (2009) Hepatobiliary fascioliasis: imaging characteristics with a new finding. Diagnostic and Interventional Radiology 15, 247.Google ScholarPubMed
Kolaskar, A and Tongaonkar, PC (1990) A semi-empirical method for prediction of antigenic determinants on protein antigens. FEBS Letters 276, 172–174.CrossRefGoogle ScholarPubMed
Lata, KS, Kumar, S, Vaghasia, V, Sharma, P, Bhairappanvar, SB, Soni, S and Das, J (2018) Exploring Leptospiral proteomes to identify potential candidates for vaccine design against Leptospirosis using an immunoinformatics approach. Scientific Reports 8, 6935.CrossRefGoogle ScholarPubMed
Levin, JM, Robson, B and Garnier, J (1986) An algorithm for secondary structure determination in proteins based on sequence similarity. FEBS Letters 205, 303–308.CrossRefGoogle ScholarPubMed
Lim, JH, Mairiang, E and Ahn, GH (2008) Biliary parasitic diseases including clonorchiasis, opisthorchiasis and fascioliasis. Abdominal Imaging 33, 157.CrossRefGoogle ScholarPubMed
Losada, S, Chacon, N, Colmenares, C, Bermudez, H, Lorenzo, A, Pointier, J, Theron, A, de Noya, BA and Noya, O (2005) Schistosoma: cross-reactivity and antigenic community among different species. Experimental Parasitology 111, 182–190.CrossRefGoogle ScholarPubMed
Machicado, C, Machicado, JD, Maco, V, Terashima, A and Marcos, LA (2016) Association of Fasciola hepatica infection with liver fibrosis, cirrhosis, and cancer: a systematic review. PLoS Neglected Tropical Diseases 10, e0004962.CrossRefGoogle ScholarPubMed
Martínez-Sernández, V, Muiño, L, Perteguer, MJ, et al. (2011) Development and evaluation of a new lateral flow immunoassay for serodiagnosis of human fasciolosis. PLoS Neglected Tropical Diseases 5, e1376.CrossRefGoogle ScholarPubMed
Mas-Coma, S, Bargues, M and Valero, M (2014) Diagnosis of human fascioliasis by stool and blood techniques: update for the present global scenario. Parasitology 141, 1918–1946.CrossRefGoogle ScholarPubMed
Miles, S, Navatta, M, Dematteis, S and Mourglia-Ettlin, G (2017) Identification of universal diagnostic peptide candidates for neglected tropical diseases caused by cestodes through the integration of multi-genome-wide analyses and immunoinformatic predictions. Infection, Genetics and Evolution 54, 338–346.CrossRefGoogle ScholarPubMed
Miles, S, Portela, M, Cyrklaff, M, Ancarola, ME, Frischknecht, F, Durán, R, Dematteis, S and Mourglia-Ettlin, G (2019) Combining proteomics and bioinformatics to explore novel tegumental antigens as vaccine candidates against Echinococcus granulosus infection. Journal of Cellular Biochemistry 120, 15320–15336.CrossRefGoogle ScholarPubMed
Mokhtarian, K, Akhlaghi, L, Meamar, AR, Razmjou, E, Naeini, KM, Gholami, S, Samei, MN and Falak, R (2016) Serodiagnosis of fasciolosis by fast protein liquid chromatography-fractionated excretory/secretory antigens. Parasitology Research 115, 2957–2965.CrossRefGoogle ScholarPubMed
Nazar, E, Azmoudeh-Ardalan, F, Mobedi, I, Dashti, SH, Salahshour, F and Ahmadinejad, Z (2017) Fasciola hepatica in explanted cirrhotic liver of a patient with chronic hepatitis B virus infection: an accidental finding. Hepatitis Monthly 17, e61214.CrossRefGoogle Scholar
O'Neill, SM, Parkinson, M, Strauss, W, Angles, R and Dalton, JP (1998) Immunodiagnosis of Fasciola hepatica infection (fascioliasis) in a human population in the Bolivian Altiplano using purified cathepsin L cysteine proteinase. The American Journal of Tropical Medicine and Hygiene 58, 417–423.CrossRefGoogle Scholar
Pan, W, Chen, DS, Lu, YJ, et al. (2017) Bioinformatic prediction of the epitopes of Echinococcus granulosus antigen 5. Biomedical Reports 6, 181–187.CrossRefGoogle ScholarPubMed
Parker, J, Guo, D and Hodges, R (1986) New hydrophilicity scale derived from high-performance liquid chromatography peptide retention data: correlation of predicted surface residues with antigenicity and X-ray-derived accessible sites. Biochemistry 25, 5425–5432.CrossRefGoogle ScholarPubMed
Pellequer, J-L, Westhof, E and Van Regenmortel, MH (1993) Correlation between the location of antigenic sites and the prediction of turns in proteins. Immunology Letters 36, 83–99.CrossRefGoogle ScholarPubMed
Pettersen, EF, Goddard, TD, Huang, CC, Couch, GS, Greenblatt, DM, Meng, EC and Ferrin, TE (2004) UCSF Chimera—a visualization system for exploratory research and analysis. Journal of computational chemistry 25, 1605–1612.CrossRefGoogle ScholarPubMed
Rohani, S, Hajighasemi, F and Sefid, F (2018) Identification of conformational epitopes on fragment crystallizable region of human Immunoglobulin G by immunoinformatic. Tehran University Medical Journal TUMS Publications 76, 321–325.Google Scholar
Rojas-Caraballo, J, López-Abán, J, Del Villar, LP, Vizcaíno, C, Vicente, B, Fernández-Soto, P, Del Olmo, E, Patarroyo, MA and Muro, A (2014) In vitro and in vivo studies for assessing the immune response and protection-inducing ability conferred by Fasciola hepatica-derived synthetic peptides containing B-and T-cell epitopes. PloS One 9, e105323.CrossRefGoogle ScholarPubMed
Rojas-Caraballo, J, López-Abán, J, Moreno-Pérez, DA, Vicente, B, Fernández-Soto, P, Del Olmo, E, Patarroyo, MA and Muro, A (2017) Transcriptome profiling of gene expression during immunisation trial against Fasciola hepatica: identification of genes and pathways involved in conferring immunoprotection in a murine model. BMC Infectious Diseases 17, 94.CrossRefGoogle Scholar
Rokni, M and Gharavi, M (2002) Evaluation of a pre-selected epitope of Fasciola hepatica cathepsin-L1 for the diagnosis of human fasciolosis by IgG-ELISA test. Iranian Journal of Public Health 31, 80–82.Google Scholar
Sabry, M, Taher, E, Farag Allah, N and Mahgoub, A (2014) Diagnosis of Fasciola infection by SDS–PAGE eluted excretory secretory (ES) protein fractions using dot-ELISA. International Journal of Veterinary Science and Medicine 2, 130–135.CrossRefGoogle Scholar
Saha, S and Raghava, GPS (2006) Prediction of continuous B-cell epitopes in an antigen using recurrent neural network. Proteins: Structure, Function, and Bioinformatics 65, 40–48.CrossRefGoogle Scholar
Salimi-Bejestani, M, McGarry, J, Felstead, S, Ortiz, P, Akca, A and Williams, DJ (2005) Development of an antibody-detection ELISA for Fasciola hepatica and its evaluation against a commercially available test. Research in Veterinary Science 78, 177–181.CrossRefGoogle ScholarPubMed
Santana, BG, Dalton, JP, Camargo, FV, Parkinson, M and Ndao, M (2013) The diagnosis of human fascioliasis by enzyme-linked immunosorbent assay (ELISA) using recombinant cathepsin L protease. PLoS Neglected Tropical Diseases 7, e2414.Google Scholar
Sarkari, B and Khabisi, SA (2017) Immunodiagnosis of human fascioliasis: an update of concepts and performances of the serological assays. Journal of Clinical and Diagnostic Research: JCDR 11, OE05.Google ScholarPubMed
Shafiei, R, Sarkari, B and Sadjjadi, SM (2015) Performance of a 27 kDa Fasciola hepatica antigen in the diagnosis of human fascioliasis. Journal of Laboratory Physicians 7, 17.Google Scholar
Singh, H, Ansari, HR and Raghava, GP (2013) Improved method for linear B-cell epitope prediction using antigen's primary sequence. PloS One 8, e62216.Google ScholarPubMed
Torres, D and Espino, AM (2006) Mapping of B-cell epitopes on a novel 11.5-kilodalton Fasciola hepatica-Schistosoma mansoni cross-reactive antigen belonging to a member of the F. hepatica saposin-like protein family. Infection and Immunity 74, 4932–4938.CrossRefGoogle Scholar
Valero, MA, Perez-Crespo, I, Periago, MV, Khoubbane, M and Mas-Coma, S (2009) Fluke egg characteristics for the diagnosis of human and animal fascioliasis by Fasciola hepatica and F. gigantica. Acta Tropica 111, 150–159.CrossRefGoogle Scholar
Valero, MA, Periago, MV, Pérez-Crespo, I, et al. (2012) Field evaluation of a coproantigen detection test for fascioliasis diagnosis and surveillance in human hyperendemic areas of Andean countries. PLoS Neglected Tropical Diseases 6, e1812.CrossRefGoogle ScholarPubMed
Vázquez-Talavera, J, Solís, CF, Medina-Escutia, E, López, ZM, Proaño, J, Correa, D and Laclette, JP (2001) Human T and B cell epitope mapping of Taenia solium paramyosin. Parasite Immunology 23, 575–579.CrossRefGoogle Scholar
Wang, Y, Wang, G and Cai, JP (2016) Identifying novel B cell epitopes within Toxoplasma gondii GRA6. The Korean Journal of Parasitology 54, 431.CrossRefGoogle Scholar
Yamaguti, S (1970) Digenetic Trematodes of Hawaiian Fishes, 436 pp. Tokyo, Japan: Keigaku Publishing Co.Google Scholar
Yang, J and Zhang, Y (2015) I-TASSER server: new development for protein structure and function predictions. Nucleic Acids Research 43, W174–W181.CrossRefGoogle ScholarPubMed
Zimic, M, Gutiérrez, AH, Gilman, RH, López, C, Quiliano, M, Evangelista, W, Gonzales, A, García, HH and Sheen, P (2011) Immunoinformatics prediction of linear epitopes from Taenia solium TSOL18. Bioinformation 6, 271.CrossRefGoogle ScholarPubMed