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PMID: 1531368 Published · ppublish English Journal Article

Cleavage of the dengue virus polyprotein at the NS3/NS4A and NS4B/NS5 junctions is mediated by viral protease NS2B-NS3, whereas NS4A/NS4B may be processed by a cellular protease.

Journal of virology ·Vol. 66 ·No. 3 ·1992-03-00 ·Pages 1535-42

Cahour A, Falgout B, Lai CJ

Abstract

The cleavage mechanism utilized for processing of the NS3-NS4A-NS4B-NS5 domain of the dengue virus polyprotein was studied by using the vaccinia virus expression system. Recombinant vaccinia viruses vNS2B-NS3-NS4A-NS4B-NS5, vNS3-NS4A-NS4B-NS5, vNS4A-NS4B-NS5, and vNS4B-NS5 were constructed. These recombinants were used to infect cells, and the labeled lysates were analyzed by immunoprecipitation. Recombinant vNS2B-NS3-NS4A-NS4B-NS5 expressed the authentic NS3 and NS5 proteins, but the other recombinants produced uncleaved polyproteins. These findings indicate that NS2B is required for processing of the downstream nonstructural proteins, including the NS3/NS4A and NS4B/NS5 junctions, both of which contain a dibasic amino acid sequence preceding the cleavage site. The flavivirus NS4A/NS4B cleavage site follows a long hydrophobic sequence. The polyprotein NS4A-NS4B-NS5 was cleaved at the NS4A/NS4B junction in the absence of other dengue virus functions. One interpretation for this finding is that NS4A/NS4B cleavage is mediated by a host protease, presumably a signal peptidase. Although vNS3-NS4A-NS4B-NS5 expressed only the polyprotein, earlier results demonstrated that cleavage at the NS4A/NS4B junction occurred when an analogous recombinant, vNS3-NS4A-84%NS4B, was expressed. Thus, it appears that uncleaved NS3 plus NS5 inhibit NS4A/NS4B cleavage presumably because the putative signal sequence is not accessible for recognition by the responsible protease. Finally, recombinants that expressed an uncleaved NS4B-NS5 polyprotein, such as vNS4A-NS4B-NS5 or vNS4B-NS5, produced NS5 when complemented with vNS2B-30%NS3 or with vNS2B plus v30%NS3. These results indicate that cleavage at the NS4B/NS5 junction can be mediated by NS2B and NS3 in trans.

MeSH Terms
Animals Base Sequence Capsid/metabolism Cells, Cultured Chlorocebus aethiops Dengue Virus/enzymology,metabolism Endopeptidases/metabolism Molecular Sequence Data Oligodeoxyribonucleotides/chemistry Polymerase Chain Reaction Protein Processing, Post-Translational Proteins/metabolism Recombinant Fusion Proteins/metabolism Viral Core Proteins/metabolism Viral Nonstructural Proteins
Chemicals
Oligodeoxyribonucleotides Proteins Recombinant Fusion Proteins Viral Core Proteins Viral Nonstructural Proteins Endopeptidases
Authors & Affiliations
3 authors, click to expand affiliations / ORCID
Cahour A
Molecular Viral Biology Section, National Institute of Allergy and Infectious Diseases, Bethesda, Maryland 20892.
Falgout B
Lai C J
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45 references, click to expand
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Article Info
Journal
Journal of virology
Abbr.
J Virol
ISSN
0022-538X
DOI
10.1128/JVI.66.3.1535-1542.1992
Published
1992-03-00
Pages
1535-42
Language
English
Region
United States
NLM ID
0113724
PMCID
PMC240879
Subset
IM
External Links
PubMed source DOI: 10.1128/JVI.66.3.1535-1542.1992 PMC Full Text View journal details
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