Super induction of dengue virus NS1 protein in E. coli.
The non-structural protein 1 (NS1) of dengue virus is a useful target for diagnostics of dengue infection since the protein is abundantly circulating in blood during the acute phase of the disease. Prior work has established that secreted NS1 levels in plasma correlates with viremia levels and hence can also be used to diagnose patients at the risk for developing dengue hemorrhagic fever. Thus detection of non-structural dengue antigens may be of benefit for an early rapid diagnosis of dengue infection due to its long half life in the blood. Here we describe a simple and efficient method for the expression of NS1 in Escherichia coli, which could potentially be used to develop monoclonal and bispecific antibodies for point of care diagnostics. E. coli codon optimized synthetic full-length NS1 gene of dengue serotype 1 (DEN-1) was successfully cloned and expressed in very high-level as inclusion bodies. The NS1 protein was successfully affinity purified and refolded as a recombinant NS1 (rNS1) protein in E. coli and yield was 230-250 mg/L of bacterial culture. The rNS1 protein was used to immunize mice for hybridoma development. The polyclonal antiserum from animals immunized with this rNS1 protein was found to specifically recognize the rNS1, thus demonstrating the immunogenic nature of the protein. The rNS1 protein purified from E. coli could be useful for developing a sensitive serum diagnostic assay to monitor dengue outbreaks.
Das
Dipankar
D
Faculty of Pharmacy and Pharmaceutical Sciences, University of Alberta, 11304-89 Avenue, Edmonton, Alberta, Canada.
Mongkolaungkoon
Sunundha
S
Suresh
Mavanur R
MR
D000818:Animals; D002648:Child; D003715:Dengue / Q000175:diagnosis* / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D002479:Inclusion Bodies / Q000378:metabolism; D051379:Mice; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism*; D017361:Viral Nonstructural Proteins / Q000235:genetics / Q000378:metabolism*; D055815:Young Adult
C082515:NS1 protein, dengue-1 virus; D011994:Recombinant Proteins; D017361:Viral Nonstructural Proteins