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Sains Malaysiana 54(5)(2025): 1405-1416
http://doi.org/10.17576/jsm-2025-5405-16
(Sebatian
Terbitan Stirilpiron (SPD), Perencat NS3, NS5 dan prM DENV-2 Novel; Merencat
Replikasi RNA, Transkripsi Protein dan Penghasilan Zarah Progeni DENV-2)
NOOR ZARINA ABD WAHAB1,* & NAZLINA IBRAHIM2
1School of Biomedicine, Faculty of Health Sciences, Universiti Sultan Zainal Abidin, 21300
Kuala Nerus, Terengganu, Malaysia
2Department of Biological Sciences and Biotechnology, Faculty
of Science and Technology, Universiti Kebangsaan Malaysia, 43600 UKM Bangi, Selangor, Malaysia
Received: 29 April
2024/Accepted: 3 January 2025
Abstract
In the last few decades, dengue has become much more
commonplace worldwide, and it is quickly spreading to other countries. As of
right now, supportive care is the only available treatment for dengue infection
as there are no approved antivirals. Although a DENV vaccine has recently been
used in some countries, its indication is limited due to risk of severe dengue
in certain individu. The present study was aimed to investigate
the effects of styrylpyrone derivative (SPD)
extracted from Goniothalamus umbrosus against DENV-2 prM,
NS3, and NS5 genes and proteins. Gene expression analysis by qRT-PCR was done to investigate the level of gene
expression at different DENV-2’s replication phases. In situ ELISA
assay was used to evaluate the effects of SPD treatment on the protein
expression of NS3, NS5, and prM. In silico molecular docking was used to understand the interactions between peptide with
target proteins. SPD inhibited the formation of infectious mature virus
particles in the process of DENV-2 replication cycle by modifying the
expression of prM gene during the infection. In
situ ELISA assay of infected cells confirmed that SPD inhibited prM, NS3, and NS5 proteins. Thus, the SPD has the ability
to alter DENV-2 replication cycle at the replication phase and during the
formation of infectious mature virus particles stage
and reduce progeny infectivity. Molecular docking in silico analysis
confirmed that SPD can interact with all selected virus proteins through
hydrogen bonds and other interactions. This study proved that SPD has the potential
as anti DENV-2 by interruption expression of DENV-2 NS3, NS5 and prM genes and proteins at RNA replication, protein
transcription, virus particles maturation and progeny infectivity phases of
viral replication cycle.
Keywords: DENV-2; docking; inhibitors; NS3; NS5; prM
Abstrak
Dalam beberapa dekad yang lalu, denggi telah menjadi penyakit yang biasa berlaku di seluruh dunia dan ia mudah merebak. Setakat ini, penjagaan sokongan adalah satu-satunya rawatan yang tersedia untuk merawat jangkitan denggi kerana ketiadaan agent
antivirus yang diluluskan. Walaupun setakat ini beberapa negara mula menggunakan vaksin DENV, namun kebolehgunaannya adalah terhad kerana sesetengah individu berisiko mendapat denggi yang teruk. Kajian ini bertujuan untuk menentukan kesan SPD yang diekstrak daripada Goniothalamus umbrosusterhadap gen dan protein prM, NS3
dan NS5 virus DENV-2. Analisis pengekspresan gen menggunakan qRT-PCR dilakukan untuk melihat tahap pengekspresan gen pada fasa replikasi DENV-2 yang berbeza. Ujian ELISA in situ digunakan untuk melihat kesan rawatan SPD terhadap pengekspresan protein NS3, NS5 dan prM. Pendokan molekul in
silico digunakan untuk memahami interaksi antara peptida dengan protein sasaran. SPD menghalang pembentukan zarah virus matang berdaya jangkit dalam proses kitaran replikasi DENV-2 dengan mengubah suai pengekspresan gen prM semasa jangkitan. Ujian ELISA in situ bagi sel yang dijangkiti telah mengesahkan bahawa SPD merencat protein prM, NS3 dan
NS5. Oleh itu, SPD mempunyai keupayaan untuk mengubah kitaran replikasi DENV-2 pada fasa replikasi dan semasa pembentukan peringkat zarah virus matang berdaya jangkit dan mengurangkan kebolehjangkitan progeni. Pendokan molekul in silico mengesahkan bahawa SPD boleh berinteraksi dengan semua protein virus terpilih melalui ikatan hidrogen dan interaksi-interaksi lain. Kajian ini menunjukkan bahawa SPD mempunyai potensi sebagai agen anti-DENV-2 dengan mengganggu pengekspresan gen dan protein DENV-2 NS3, NS5 dan prM dalam kitaran replikasi virus ketika replikasi RNA, transkripsi protein, pematangan zarah virus dan kebolehjangkitan progeni.
Kata kunci: DENV-2; NS3; NS5; pendokan; perencat; prM
REFERENCES
Abd Wahab, N.Z. & Ibrahim, N.
2022. Styrylpyrone derivative
(SPD) extracted from Goniothalamus umbrosus binds to dengue virus serotype-2 envelope
protein and inhibits early stage of virus replication. Molecules 27(14): 4566.
Abd Wahab, N.Z. & Ibrahim, N.
2020a. Efficacy of Catharanthus
roseus extract against dengue virus type 2 infection in vitro. Indian
Journal of Public Health Research & Development 11(1): 1320-1325.
Abd Wahab, N.Z. & Ibrahim, N. 2020b. In vitro study,
antiviral activity of styrylpyrone derivative against
dengue virus type 2. Asian Journal of Plant Sciences 19: 438-442.
Abd Wahab, N.Z.,
Ibrahim, N., Kamarudin, M.K.A. & Ghazali, A. 2018. In vitro antiviral
activity of Orthosiphon stamineus extract
against dengue virus type 2. Malaysian Journal of Fundamental and Applied
Sciences 10(1S): 541-551.
Abdullah, Z.L., Chee,
H.Y., Yusof, R. & Mohd Fauzi, F. 2023. Finding lead compounds for dengue
antivirals from a collection of old drugs through in silico target
prediction and subsequent in vitro validation. ACS Omega 8(36): 32483-32497.
Acosta, E.G., Talarico, L.B. & Damonte, E.B. 2008. Cell
entry of dengue virus. Future Virology 3(5): 471-479.
Alagarasu, K., Patil,
P., Kaushik, M., Chowdhury, D., Joshi, R.K., Hegde, H.V., Kakade, M.B., Hoti,
S.L., Cherian, S. & Parashar, D. 2022. In vitro antiviral activity
of potential medicinal plant extracts against dengue and chikungunya viruses. Frontiers
in Cellular and Infection Microbiology 12: 866452.
Alves, R.P.D.S., Pereira, L.R.,
Fabris, D.L.N., Salvador, F.S., Santos, R.A., Zanotto, P.M.A., Romano, C.M.,
Amorim, J.H. & Ferreira, L.C.S. 2016. Production of a recombinant dengue
virus 2 NS5 protein and potential use as a vaccine antigen. Clinical and
Vaccine Immunology 23(6): 460-469.
Antila, H., Autio,
H., Turunen, L., Harju, K., Tammela, P., Wennerberg, K., Yli-Kauhaluoma,
J., Huttunen, H.J., Castrén,
E. & Rantamäki, T. 2014. Utilization of in situ ELISA method
for examining Trk receptor phosphorylation in cultured cells. Journal of
Neuroscience Methods 222: 142-146.
Arthur, D.E. & Uzairu, A. 2019. Molecular docking studies on the
interaction of NCI anticancer analogues with human phosphatidylinositol
4,5-bisphosphate 3-kinase catalytic subunit. Journal of King Saud University
- Science 31(4): 1151-1166.
Berg, J.M., Tymoczko, J.L. & Stryer, L.
2002. Biochemistry. 5th ed. New York: W.H. Freeman.
Bernaldez, M.J.A.,
Billones, J.B. & Magpantay, A. 2018. In silico analysis of binding
interactions between GSK983 and human DHODH through docking and molecular
dynamics. AIP Conference Proceedings 2045: 020073.
Chambers,
T.J., Hahn, C.S., Galler, R. & Rice, C.M. 1990. Flavivirus genome
organization, expression, and replication. Annual Review of Microbiology 44:
649-688.
Fernandes, P.O., Chagas,
M.A., Rocha, W.R. & Moraes, A.H. 2021. Non-structural protein 5 (NS5) as a
target for antiviral development against established and emergent flaviviruses. Current Opinion in Virology 50: 30-39.
Gamez, P. 2014. The
anion–π interaction: Naissance and establishment of a peculiar
supramolecular bond. Inorganic Chemistry Frontiers 1(1): 35-43.
Iberahim, R., Md.
Nor, N.S., Yaacob, W.A. & Ibrahim, N. 2018. Eleusine indica inhibits
early and late phases of herpes simplex virus type 1 replication cycle and
reduces progeny infectivity. Sains Malaysiana47(7): 1431-1438.
Issur, M., Geiss, B.J., Bougie, I., Picard-Jean, F., Despins,
S., Mayette, J., Hobdey, S.E. & Bisaillon, M. 2009. The flavivirus NS5 protein is a true
RNA guanylyltransferase that catalyzes a two-step
reaction to form the RNA cap structure. RNA 15(12): 2340-2350.
Jayasekara, K.G., Suresh, S., Goonasekara, C., Soyza, P., Perera N.
& Gunasekera, K. 2024. Anti-dengue viral activity of Glycyrrhiza glabra roots in Vero cells. Scientific Reports 14(1): 25922.
Johansson, M., Brooks,
A.J., Jans, D.A. & Vasudevan, S.G. 2001. A small region of the dengue
virus-encoded RNA-dependent RNA polymerase, NS5, confers interaction with both
the nuclear transport receptor importin-beta and the viral helicase, NS3. Journal
of Virology 82(Pt 4): 735-745.
Junjhon, J., Edwards, T.J., Utaipat,
U., Bowman, V.D., Holdaway, H.A., Zhang, W., Keelapang,
P., Puttikhunt, C., Perera, R., Chipman, P.R., Kasinrerk, W., Malasit, P., Kuhn,
R.J. & Sittisombut, N. 2010. Influence of pr-M
cleavage on the heterogeneity of extracellular dengue virus particles. Journal
of Virology 84(16): 8353-8358.
Kapoor, M., Zhang,
L., Ramachandra, M., Kusukawa, J., Ebner, K.E. &
Padmanabhan, R. 1995. Association between NS3 and NS5 proteins of dengue virus
type 2 in the putative RNA replicase is linked to differential phosphorylation
of NS5. Journal of Biological Chemistry 270(32): 19100-19106.
Krol, E., Brzuska, G. & Szewczyk, B. 2019.
Production and biomedical application of flavivirus-like particles. Trends
in Biotechnology 37(11): 1202-1216.
Li, L., Lok,
S.M., Yu, I.M., Zhang, Y., Kuhn, R.J., Chen, J. & Rossmann, M.G. 2008. The
flavivirus precursor membrane-envelope protein complex: Structure and
maturation. Science 319(5871): 1830-1834.
Li, Q. &
Kang, C. 2022. Dengue virus NS4B protein as a target for developing
antivirals. Frontiers in Cellular and Infection Microbiology 12: 959727.
Matusan, A.E.,
Kelley, P.G., Pryor, M.J., Whisstock, J.C., Davidson,
A.D. & Wright, P.J. 2001. Mutagenesis of the dengue virus type 2 NS3
proteinase and the production of growth-restricted virus. Journal of General
Virology 82(Pt 7): 1647-1656.
Mohamad Yussoff, M.A., Abd
Hamid, A.A., Abd Hamid, S. & Abd Halim, K.B. 2020. Computational quest for
finding potential Ebola VP40 inhibitors: A molecular docking study. Sains Malaysiana 49(3): 537-544.
Mukhopadhyay, S., Kuhn,
R.J. & Rossmann, M.G. 2005. A structural perspective of the flavivirus life
cycle. Nature Reviews Microbiology 3(1): 13-22.
Murugesan, A.
& Manoharan, M. 2020. Dengue virus. Emerging Infectious Diseases 2020:
281-359.
Obi, J.O.,
Gutiérrez-Barbosa, H., Chua, J.V. & Deredge, D.J. 2021.
Current trends and limitations in dengue antiviral research. Tropical
Medicine and Infectious Disease 6(4): 180.
Othman, R., Othman,
R., Baharuddin, A., Ramakrishnan, N.R., Abd Rahman, N., Yusof, R. & Karsani, S.A. 2017. Molecular docking studies of selected
medicinal drugs as dengue virus-2 protease inhibitors. Sains Malaysiana46(10): 1865-1875.
Perera, R.
& Kuhn, R.J. 2008. Structural proteomics of dengue virus. Current
Opinion in Microbiology 11(4): 369-377.
Rajapakse, S., de Silva, N.L., Weeratunga, P., Rodrigo, C., Sigera,
C. & Fernando, S.D. 2019. Carica
papaya extract in dengue: A systematic review and meta-analysis. BMC
Complementary and Alternative Medicine 9(1): 265.
Ribas, J., Cubero, E., Luque, F.J. & Orozco, M. 2002.
Theoretical study of alkyl-pi and aryl-pi interactions. Reconciling theory and
experiment. Journal of Organic Chemistry 67(20): 7057-7065.
Rodenhuis-Zybert, I.A., van
der Schaar, H.M., da Silva Voorham, J.M., van der
Ende-Metselaar, H., Lei, H.Y., Wilschut,
J. & Smit, J.M. 2010. Immature dengue virus: A veiled pathogen? PLOS
Pathogens 6(1): e1000718.
Rothan, H.A., Han, H.C., Ramasamy, T.S., Othman, S., Rahman,
N.A. & Yusof, R. 2012. Inhibition of dengue NS2B-NS3 protease and viral
replication in Vero cells by recombinant retrocyclin-1. BMC Infectious
Diseases 12: 314.
Saleem, H.N.,
Batool, F., Mansoor, H.J., Shahzad-ul-Hussan, S.
& Saeed, M. 2019. Inhibition of dengue virus protease by eugeniin, isobiflorin, and biflorin isolated from the flower buds of Syzygium aromaticum (Cloves). ACS Omega 4(1): 1525-1533.
Shimizu, H., Saito,
A., Mikuni, J., Nakayama, E.E., Koyama, H., Honma, T., Shirouzu, M., Sekine,
S.I. & Shioda, T. 2019. Discovery of a small molecule inhibitor targeting
dengue virus NS5 RNA-dependent RNA polymerase. PLOS Neglected Tropical
Diseases 13(11): e0007894.
Sinha, S., Singh, K., Ravi Kumar, Y.S., Roy, R., Phadnis, S.,
Meena, V., Bhattacharyya, S. & Verma, B. 2024. Dengue virus pathogenesis
and host molecular machineries. Journal
of Biomedical Science 31(1): 43.
Stolp, Z.D.,
Smurthwaite, C.A., Reed, C., Williams, W., Dharmawan,
A., Djaballah, H. & Wolkowicz, R. 2015. A multiplexed cell-based assay for
the identification of modulators of pre-membrane processing as a target against
dengue virus. Journal of Biomolecular Screening 20(5): 616-626.
Tay, M.Y., Saw, W.G., Zhao, Y., Chan, K.W., Singh, D.,
Chong, Y., Forwood, J.K., Ooi, E.E., Grber, G., Lescar, J., Luo, D.
& Vasudevan, S.G. 2015. The C-terminal 50 amino acid residues of dengue NS3
protein are important for NS3-NS5 interaction and viral replication. Journal
of Biological Chemistry 290(4): 2379-2394.
van den Elsen, K., Quek, J.P. & Luo, D. 2021. Molecular insights
into the flavivirus replication complex. Viruses 13(6): 956.
WHO. 2024. https://www.who.int/news-room/fact-sheets/detail/dengue-and-severe-dengue
Wu, Y.S., Afzal, S., Appalaraju, V., Wei, T.Q., Abdul Manap, A.S. & Albokhadaim, I. 2024. Evaluation for antiviral potential of Ficus deltoidea against dengue virus type-2. Sains Malaysiana 53(2): 383-396.
Yahya, A.K., Abd Wahab, N.Z. &
Ibrahim, N. 2024. Bioactive compounds of plant essential oils and their
antiviral properties: A comprehensive review. Malaysian Journal of Chemistry 26(4): 123-136.
Yon, C., Teramoto, T., Mueller, N., Phelan, J., Ganesh,
V.K., Murthy, K.H. & Padmanabhan, R. 2005. Modulation of the nucleoside
triphosphatase/RNA helicase and 5'-RNA triphosphatase activities of Dengue
virus type 2 nonstructural protein 3 (NS3) by interaction with NS5, the
RNA-dependent RNA polymerase. Journal of Biological Chemistry 280(29): 27412-27419.
Zhang, T., Wang,
M.L., Zhang, G.R., Liu, W., Xiao, X.Q., Yang, Y.S., Li, J.T., Xun, Z.M., Li,
D.Y. & Chan, P.K.S. 2019. Recombinant DENV 2 NS5: An effective antigen for
diagnosis of DENV infection. Journal of Virological Methods 265: 35-41.
Zhang, X., Zhang, Y., Jia, R.,
Wang, M., Yin, Z. & Cheng, A. 2021. Structure and function of capsid
protein in flavivirus infection and its applications in the development of
vaccines and therapeutics. Veterinary Research 52(1): 98.
Zheng, A., Umashankar, M. & Kielian, M. 2010. In
vitro and in vivo studies identify important features of
dengue virus pr-E protein interactions. PLOS Pathogens 6(10): e1001157.
*Corresponding author;
email: zarinawahab@unisza.edu.my
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