The Bioinformatics Analysis and Single-chain Fragment Variable Expression for Detection of Indonesian Dengue Virus NS1 Antigen
Keywords:
dengue virus, molecular docking, molecular dynamics simulation, NS1 antigen, scFv
Abstract
Dengue hemorrhagic fever is a major global health challenge, the effective management of which requires early and accurate detection. Nonstructural protein 1 (NS1) has been shown to be a potential biomarker which offers a diagnostic window from the first day of infection. This study aims to explore the diagnostic potential of a modified antibody engineered as a single-chain variable fragment (scFv) for specific NS1 detection in DHF cases in Indonesia. The interactions between scFv and NS1 DENV Indonesia were analyzed using computational approaches, which include molecular docking and molecular dynamics simulations. The successful docking and subsequent enzyme-linked immunosorbent assay experiments confirmed the binding of the scFv to NS1. In silico approaches complemented experimental data, which provided valuable insights into potential scFv-NS1 interactions and guided targeted diagnostic tool development. The use of scFv as a specific tool has the potential to overcome the current limitations in sensitivity observed in NS1 detection in Indonesia. In this study, the modified scFv was produced in Escherichia coli BL21(DE3) host recombinantly, hence offering advantages over conventional antibodies. To conclude, our study offers a promising avenue for early and specific NS1 detection in DHF cases using modified scFv. The combination of computational analysis with experimental validation underscores the significance of a multidisciplinary approach in developing innovative diagnostic tools. To establish the clinical utility of scFv as a reliable and cost-effective diagnostic tool for DHF, further investigations and clinical assessments are necessary. This advancement may have far-reaching implications in the fight against dengue and other infectious diseases.
References
Andries, A. C., Duong, V., Ong, S., Ros, S., Sakuntabhai, A., Horwood, P., Dussart, P., & Buchy, P. (2016). Evaluation of the performances of six commercial kits designed for dengue NS1 and anti-dengue IgM, IgG and IgA detection in urine and saliva clinical specimens. BMC Infectious Diseases, 16(1), 1–9. https://doi.org/10.1186/s12879-016-1551-x
Aryati, A., Trimarsanto, H., Yohan, B., Wardhani, P., Fahri, S., & Sasmono, R. T. (2013). Performance of commercial dengue NS1 ELISA and molecular analysis of NS1 gene of dengue viruses obtained during surveillance in Indonesia. BMC Infectious Diseases, 13(611), 1–11. https://doi.org/10.1186/1471-2334-13-611
Astriany, D., Indah, Y., Sari, P., Baroroh, U., Yusuf, M., Maksum, I. P., Natalia, D., & Subroto, T. (2020). In Silico Study of Single Chain Fragment Variable Antibody and Indonesian Serotype-2 NS1 Dengue Virus Antigen. Advances in Health Sciences Research, 26, 102–106. http://rosettaserver.graylab.jhu.edu/
Buchman, A., Gamez, S., Li, M., Antoshechkin, I., Li, H. H., Wang, H. W., Chen, C. H., Klein, M. J., Duchemin, J. B., Crowe, J. E., Paradkar, P. N., & Akbari, O. S. (2020). Broad dengue neutralization in mosquitoes expressing an engineered antibody. PLoS Pathogens, 16(1), 1–20. https://doi.org/10.1371/journal.ppat.1008103
Chen, P. K., Chang, J. H., Ke, L. Y., Kao, J. K., Chen, C. H., Yang, R. C., Yoshimura, T., Ito, E., & Tsai, J. J. (2023). Advanced Detection Method for Dengue NS1 Protein Using Ultrasensitive ELISA with Thio-NAD Cycling. Viruses, 15(1894), 1–8. https://doi.org/10.3390/v15091894
Crisan, L., Funar-Timofei, S., & Borota, A. (2022). Homology Modeling and Molecular Docking Approaches for the Proposal of Novel Insecticides against the African Malaria Mosquito (Anopheles gambiae). Molecules, 27(3846), 1–14. https://doi.org/10.3390/molecules27123846
Falzone, L., Gattuso, G., Tsatsakis, A., Spandidos, D. A., & Libra, M. (2021). Current and innovative methods for the diagnosis of COVID-19 infection (Review). International Journal of Molecular Medicine, 47(6), 1–23. https://doi.org/10.3892/ijmm.2021.4933
Gelanew, T., & Hunsperger, E. (2018). Development and characterization of serotype-specific monoclonal antibodies against the dengue virus-4 (DENV-4) non-structural protein (NS1). Virology Journal, 15(30), 1–12. https://doi.org/10.1186/s12985-018-0925-7
Harapan, H., Michie, A., Mudatsir, M., Sasmono, R. T., & Imrie, A. (2019). Epidemiology of dengue hemorrhagic fever in Indonesia: Analysis of five decades data from the National Disease Surveillance. BMC Research Notes, 12(350), 1–6. https://doi.org/10.1186/s13104-019-4379-9
Hussain, W., Qaddir, I., Mahmood, S., & Rasool, N. (2018). In silico targeting of non-structural 4B protein from dengue virus 4 with spiropyrazolopyridone: study of molecular dynamics simulation, ADMET and virtual screening. Virus Disease, 29(2), 147–156. https://doi.org/10.1007/s13337-018-0446-4
Khetarpal, N., & Khanna, I. (2016). Dengue Fever: Causes, Complications, and Vaccine Strategies. In Journal of Immunology Research (Vol. 2016, pp. 1–14). Hindawi Publishing Corporation. https://doi.org/10.1155/2016/6803098
Kordi, S., Rahmati-Yamchi, M., Vostakolaei, M. A., Barzegari, A., & Abdolalizadeh, J. (2019). Purification of a novel anti-VEGFR2 single chain antibody fragment and evaluation of binding affinity by surface plasmon resonance. Advanced Pharmaceutical Bulletin, 9(1), 64–69. https://doi.org/10.15171/apb.2019.008
Laskowski, R. A., Macarthur, M. W., & Thornton, J. M. (2012). PROCHECK: validation of protein-structure coordinates. In International Tables for Crystallography (pp. 684–687). https://onlinelibrary.wiley.com/iucr/itc/Fb/ch21o4v0001/ch21o4.pdf
Lee, H., Ryu, J. H., Park, H. S., Park, K. H., Bae, H., Yun, S., Choi, A. R., Cho, S. Y., Park, C., Lee, D. G., Lim, J., Lee, J., Lee, S., Shin, S., Park, H., & Oh, E. J. (2019). Comparison of Six Commercial Diagnostic Tests for the Detection of Dengue Virus Non-Structural-1 Antigen and IgM/IgG Antibodies. Annals of Laboratory Medicine, 39(6), 566–571. https://doi.org/10.3343/alm.2019.39.6.566
Leman, J. K., Weitzner, B. D., Lewis, S. M., Adolf-Bryfogle, J., Alam, N., Alford, R. F., Aprahamian, M., Baker, D., Barlow, K. A., Barth, P., Basanta, B., Bender, B. J., Blacklock, K., Bonet, J., Boyken, S. E., Bradley, P., Bystroff, C., Conway, P., Cooper, S., … Bonneau, R. (2020). Macromolecular modeling and design in Rosetta: recent methods and frameworks. In Nature Methods (Vol. 17, Issue 7, pp. 665–680). Nature Research. https://doi.org/10.1038/s41592-020-0848-2
Mashiach, E., Schneidman-Duhovny, D., Andrusier, N., Nussinov, R., & Wolfson, H. J. (2008). FireDock: a web server for fast interaction refinement in molecular docking. Nucleic Acids Research, 36(Web Server issue), 229–232. https://doi.org/10.1093/nar/gkn186
Ministry of Health Republic of Indonesia. (2021). Indonesian Health Profile 2020. Jakarta, Indonesia: Kementerian Kesehatan Republik Indonesia, pp. 191-195
Modhiran, N., Song, H., Liu, L., Bletchly, C., Brillault, L., Amarilla, A. A., Xu, X., Qi, J., Chai, Y., Cheung, S. T. M., Traves, R., Setoh, Y. X., Bibby, S., Scott, C. A. P., Freney, M. E., Newton, N. D., Khromykh, A. A., Chappell, K. J., Muller, D. A., … Watterson, D. (2021). A broadly protective antibody that targets the flavivirus NS1 protein. Science, 371(6525), 190–194. https://doi.org/10.1126/science.abb9425
Parkash, O., & Shueb, R. H. (2015). Diagnosis of Dengue Infection Using Conventional and Biosensor Based Techniques. Viruses, 7(September), 5410–5427. https://doi.org/10.3390/v7102877
Pereira, S. S., Andreata-Santos, R., Pereira, L. R., Soares, C. P., Félix, A. C., de Andrade, P. de M. J. C., Durigon, E. L., Romano, C. M., & Ferreira, L. C. de S. (2021). NS1-based ELISA test efficiently detects dengue infections without cross-reactivity with Zika virus. International Journal of Infectious Diseases, 112, 202–204. https://doi.org/10.1016/j.ijid.2021.09.009
Sayers, E. W., Bolton, E. E., Brister, J. R., Canese, K., Chan, J., Comeau, D. C., Connor, R., Funk, K., Kelly, C., Kim, S., Madej, T., Marchler-Bauer, A., Lanczycki, C., Lathrop, S., Lu, Z., Thibaud-Nissen, F., Murphy, T., Phan, L., Skripchenko, Y., … Sherry, S. T. (2022). Database resources of the national center for biotechnology information. Nucleic Acids Research, 50(D1), D20–D26. https://doi.org/10.1093/nar/gkab1112
Schneidman-Duhovny, D., Inbar, Y., Nussinov, R., & Wolfson, H. J. (2005). PatchDock and SymmDock: Servers for rigid and symmetric docking. Nucleic Acids Research, 33(SUPPL. 2), 363–367. https://doi.org/10.1093/nar/gki481
Songprakhon, P., Thaingtamtanha, T., Limjindaporn, T., Puttikhunt, C., Srisawat, C., Luangaram, P., Dechtawewat, T., Uthaipibull, C., Thongsima, S., Yenchitsomanus, P. thai, Malasit, P., & Noisakran, S. (2020). Peptides targeting dengue viral nonstructural protein 1 inhibit dengue virus production. Scientific Reports, 10(12933), 1–16. https://doi.org/10.1038/s41598-020-69515-9
Vora, J., Patel, S., Athar, M., Sinha, S., Chhabria, M. T., Jha, P. C., & Shrivastava, N. (2020). Pharmacophore modeling, molecular docking and molecular dynamics simulation for screening and identifying anti-dengue phytocompounds. Journal of Biomolecular Structure and Dynamics, 38(6), 1726–1740.
https://doi.org/10.1080/07391102.2019.1615002
Weitzner, B. D., Jeliazkov, J. R., Lyskov, S., Marze, N., Kuroda, D., Frick, R., Adolf-Bryfogle, J., Biswas, N., Dunbrack, R. L., & Gray, J. J. (2017). Modeling and docking of antibody structures with Rosetta. Nature Protocols, 12(2), 401–416. https://doi.org/10.1038/nprot.2016.180
Aryati, A., Trimarsanto, H., Yohan, B., Wardhani, P., Fahri, S., & Sasmono, R. T. (2013). Performance of commercial dengue NS1 ELISA and molecular analysis of NS1 gene of dengue viruses obtained during surveillance in Indonesia. BMC Infectious Diseases, 13(611), 1–11. https://doi.org/10.1186/1471-2334-13-611
Astriany, D., Indah, Y., Sari, P., Baroroh, U., Yusuf, M., Maksum, I. P., Natalia, D., & Subroto, T. (2020). In Silico Study of Single Chain Fragment Variable Antibody and Indonesian Serotype-2 NS1 Dengue Virus Antigen. Advances in Health Sciences Research, 26, 102–106. http://rosettaserver.graylab.jhu.edu/
Buchman, A., Gamez, S., Li, M., Antoshechkin, I., Li, H. H., Wang, H. W., Chen, C. H., Klein, M. J., Duchemin, J. B., Crowe, J. E., Paradkar, P. N., & Akbari, O. S. (2020). Broad dengue neutralization in mosquitoes expressing an engineered antibody. PLoS Pathogens, 16(1), 1–20. https://doi.org/10.1371/journal.ppat.1008103
Chen, P. K., Chang, J. H., Ke, L. Y., Kao, J. K., Chen, C. H., Yang, R. C., Yoshimura, T., Ito, E., & Tsai, J. J. (2023). Advanced Detection Method for Dengue NS1 Protein Using Ultrasensitive ELISA with Thio-NAD Cycling. Viruses, 15(1894), 1–8. https://doi.org/10.3390/v15091894
Crisan, L., Funar-Timofei, S., & Borota, A. (2022). Homology Modeling and Molecular Docking Approaches for the Proposal of Novel Insecticides against the African Malaria Mosquito (Anopheles gambiae). Molecules, 27(3846), 1–14. https://doi.org/10.3390/molecules27123846
Falzone, L., Gattuso, G., Tsatsakis, A., Spandidos, D. A., & Libra, M. (2021). Current and innovative methods for the diagnosis of COVID-19 infection (Review). International Journal of Molecular Medicine, 47(6), 1–23. https://doi.org/10.3892/ijmm.2021.4933
Gelanew, T., & Hunsperger, E. (2018). Development and characterization of serotype-specific monoclonal antibodies against the dengue virus-4 (DENV-4) non-structural protein (NS1). Virology Journal, 15(30), 1–12. https://doi.org/10.1186/s12985-018-0925-7
Harapan, H., Michie, A., Mudatsir, M., Sasmono, R. T., & Imrie, A. (2019). Epidemiology of dengue hemorrhagic fever in Indonesia: Analysis of five decades data from the National Disease Surveillance. BMC Research Notes, 12(350), 1–6. https://doi.org/10.1186/s13104-019-4379-9
Hussain, W., Qaddir, I., Mahmood, S., & Rasool, N. (2018). In silico targeting of non-structural 4B protein from dengue virus 4 with spiropyrazolopyridone: study of molecular dynamics simulation, ADMET and virtual screening. Virus Disease, 29(2), 147–156. https://doi.org/10.1007/s13337-018-0446-4
Khetarpal, N., & Khanna, I. (2016). Dengue Fever: Causes, Complications, and Vaccine Strategies. In Journal of Immunology Research (Vol. 2016, pp. 1–14). Hindawi Publishing Corporation. https://doi.org/10.1155/2016/6803098
Kordi, S., Rahmati-Yamchi, M., Vostakolaei, M. A., Barzegari, A., & Abdolalizadeh, J. (2019). Purification of a novel anti-VEGFR2 single chain antibody fragment and evaluation of binding affinity by surface plasmon resonance. Advanced Pharmaceutical Bulletin, 9(1), 64–69. https://doi.org/10.15171/apb.2019.008
Laskowski, R. A., Macarthur, M. W., & Thornton, J. M. (2012). PROCHECK: validation of protein-structure coordinates. In International Tables for Crystallography (pp. 684–687). https://onlinelibrary.wiley.com/iucr/itc/Fb/ch21o4v0001/ch21o4.pdf
Lee, H., Ryu, J. H., Park, H. S., Park, K. H., Bae, H., Yun, S., Choi, A. R., Cho, S. Y., Park, C., Lee, D. G., Lim, J., Lee, J., Lee, S., Shin, S., Park, H., & Oh, E. J. (2019). Comparison of Six Commercial Diagnostic Tests for the Detection of Dengue Virus Non-Structural-1 Antigen and IgM/IgG Antibodies. Annals of Laboratory Medicine, 39(6), 566–571. https://doi.org/10.3343/alm.2019.39.6.566
Leman, J. K., Weitzner, B. D., Lewis, S. M., Adolf-Bryfogle, J., Alam, N., Alford, R. F., Aprahamian, M., Baker, D., Barlow, K. A., Barth, P., Basanta, B., Bender, B. J., Blacklock, K., Bonet, J., Boyken, S. E., Bradley, P., Bystroff, C., Conway, P., Cooper, S., … Bonneau, R. (2020). Macromolecular modeling and design in Rosetta: recent methods and frameworks. In Nature Methods (Vol. 17, Issue 7, pp. 665–680). Nature Research. https://doi.org/10.1038/s41592-020-0848-2
Mashiach, E., Schneidman-Duhovny, D., Andrusier, N., Nussinov, R., & Wolfson, H. J. (2008). FireDock: a web server for fast interaction refinement in molecular docking. Nucleic Acids Research, 36(Web Server issue), 229–232. https://doi.org/10.1093/nar/gkn186
Ministry of Health Republic of Indonesia. (2021). Indonesian Health Profile 2020. Jakarta, Indonesia: Kementerian Kesehatan Republik Indonesia, pp. 191-195
Modhiran, N., Song, H., Liu, L., Bletchly, C., Brillault, L., Amarilla, A. A., Xu, X., Qi, J., Chai, Y., Cheung, S. T. M., Traves, R., Setoh, Y. X., Bibby, S., Scott, C. A. P., Freney, M. E., Newton, N. D., Khromykh, A. A., Chappell, K. J., Muller, D. A., … Watterson, D. (2021). A broadly protective antibody that targets the flavivirus NS1 protein. Science, 371(6525), 190–194. https://doi.org/10.1126/science.abb9425
Parkash, O., & Shueb, R. H. (2015). Diagnosis of Dengue Infection Using Conventional and Biosensor Based Techniques. Viruses, 7(September), 5410–5427. https://doi.org/10.3390/v7102877
Pereira, S. S., Andreata-Santos, R., Pereira, L. R., Soares, C. P., Félix, A. C., de Andrade, P. de M. J. C., Durigon, E. L., Romano, C. M., & Ferreira, L. C. de S. (2021). NS1-based ELISA test efficiently detects dengue infections without cross-reactivity with Zika virus. International Journal of Infectious Diseases, 112, 202–204. https://doi.org/10.1016/j.ijid.2021.09.009
Sayers, E. W., Bolton, E. E., Brister, J. R., Canese, K., Chan, J., Comeau, D. C., Connor, R., Funk, K., Kelly, C., Kim, S., Madej, T., Marchler-Bauer, A., Lanczycki, C., Lathrop, S., Lu, Z., Thibaud-Nissen, F., Murphy, T., Phan, L., Skripchenko, Y., … Sherry, S. T. (2022). Database resources of the national center for biotechnology information. Nucleic Acids Research, 50(D1), D20–D26. https://doi.org/10.1093/nar/gkab1112
Schneidman-Duhovny, D., Inbar, Y., Nussinov, R., & Wolfson, H. J. (2005). PatchDock and SymmDock: Servers for rigid and symmetric docking. Nucleic Acids Research, 33(SUPPL. 2), 363–367. https://doi.org/10.1093/nar/gki481
Songprakhon, P., Thaingtamtanha, T., Limjindaporn, T., Puttikhunt, C., Srisawat, C., Luangaram, P., Dechtawewat, T., Uthaipibull, C., Thongsima, S., Yenchitsomanus, P. thai, Malasit, P., & Noisakran, S. (2020). Peptides targeting dengue viral nonstructural protein 1 inhibit dengue virus production. Scientific Reports, 10(12933), 1–16. https://doi.org/10.1038/s41598-020-69515-9
Vora, J., Patel, S., Athar, M., Sinha, S., Chhabria, M. T., Jha, P. C., & Shrivastava, N. (2020). Pharmacophore modeling, molecular docking and molecular dynamics simulation for screening and identifying anti-dengue phytocompounds. Journal of Biomolecular Structure and Dynamics, 38(6), 1726–1740.
https://doi.org/10.1080/07391102.2019.1615002
Weitzner, B. D., Jeliazkov, J. R., Lyskov, S., Marze, N., Kuroda, D., Frick, R., Adolf-Bryfogle, J., Biswas, N., Dunbrack, R. L., & Gray, J. J. (2017). Modeling and docking of antibody structures with Rosetta. Nature Protocols, 12(2), 401–416. https://doi.org/10.1038/nprot.2016.180
Published
2025-10-13
How to Cite
Astriany, D., Baroroh, U., Yusuf, M., Yusuf, M., Subroto, T., Maksum, I. P., & Natalia, D. (2025). The Bioinformatics Analysis and Single-chain Fragment Variable Expression for Detection of Indonesian Dengue Virus NS1 Antigen. Indonesian Journal of Pharmacy. https://doi.org/10.22146/ijp.17929
Issue
Section
Research Article
