Fusion of protegrin-1 and plectasin to MAP30 shows significant inhibition activity against dengue virus replication.
Dengue virus (DENV) broadly disseminates in tropical and sub-tropical countries and there are no vaccine or anti-dengue drugs available. DENV outbreaks cause serious economic burden due to infection complications that requires special medical care and hospitalization. This study presents a new strategy for inexpensive production of anti-DENV peptide-fusion protein to prevent and/or treat DENV infection. Antiviral cationic peptides protegrin-1 (PG1) and plectasin (PLSN) were fused with MAP30 protein to produce recombinant antiviral peptide-fusion protein (PG1-MAP30-PLSN) as inclusion bodies in E. coli. High yield production of PG1-MAP30-PLSN protein was achieved by solubilization of inclusion bodies in alkaline buffer followed by the application of appropriate refolding techniques. Antiviral PG1-MAP30-PLSN protein considerably inhibited DENV protease (NS2B-NS3pro) with half-maximal inhibitory concentration (IC50) 0.5±0.1 μM. The real-time proliferation assay (RTCA) and the end-point proliferation assay (MTT assay) showed that the maximal-nontoxic dose of the peptide-fusion protein against Vero cells is approximately 0.67±0.2 μM. The cell-based assays showed considerable inhibition of the peptide-fusion protein against binding and proliferating stages of DENV2 into the target cells. The peptide-fusion protein protected DENV2-challeged mice with 100% of survival at the dose of 50 mg/kg. In conclusion, producing recombinant antiviral peptide-fusion protein by combining short antiviral peptide with a central protein owning similar activity could be useful to minimize the overall cost of short peptide production and take advantage of its synergistic antiviral activities.
Department of Molecular Medicine, Faculty of Medicine, University of Malaya, Kuala Lumpur, Malaysia.
Hussin A
HA
Rothan
Department of Molecular Medicine, Faculty of Medicine, University of Malaya, Kuala Lumpur, Malaysia.
Hirbod
H
Bahrani
Genetics and Molecular Biology Unit, Institute of Biological Science, Faculty of Science, University of Malaya, Kuala Lumpur, Malaysia.
Zulqarnain
Z
Mohamed
Department of Chemistry, Faculty of Science, University of Malaya, Kuala Lumpur, Malaysia.
Noorsaadah
N
Abd Rahman
Department of Molecular Medicine, Faculty of Medicine, University of Malaya, Kuala Lumpur, Malaysia.
Rohana
R
Yusof
D000595:Amino Acid Sequence; D000818:Animals; D023181:Antimicrobial Cationic Peptides / Q000235:genetics* / Q000378:metabolism; D002522:Chlorocebus aethiops; D003715:Dengue / Q000188:drug therapy* / Q000401:mortality / Q000821:virology; D003716:Dengue Virus / Q000187:drug effects / Q000201:enzymology / Q000254:growth & development; D004305:Dose-Response Relationship, Drug; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D005260:Female; D015870:Gene Expression; D002479:Inclusion Bodies / Q000737:chemistry; D008297:Male; D051379:Mice; D008813:Mice, Inbred ICR; D008969:Molecular Sequence Data; D010455:Peptides / Q000235:genetics* / Q000378:metabolism; D058849:Protein Refolding; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000235:genetics / Q000494:pharmacology*; D054790:Ribosome Inactivating Proteins, Type 2 / Q000235:genetics* / Q000378:metabolism; D012697:Serine Endopeptidases / Q000378:metabolism; D016019:Survival Analysis; D014709:Vero Cells; D017361:Viral Nonstructural Proteins / Q000037:antagonists & inhibitors / Q000378:metabolism; D000085242:Viral Proteases
D023181:Antimicrobial Cationic Peptides; D010455:Peptides; D011993:Recombinant Fusion Proteins; D054790:Ribosome Inactivating Proteins, Type 2; D012697:Serine Endopeptidases; D017361:Viral Nonstructural Proteins; C066347:MAP30 protein, Momordica charantia; C080102:NS2B protein, flavivirus; C504857:plectasin; C094880:protegrin-1; C504449:NS3 protease, dengue virus; D000085242:Viral Proteases