Antigenicity of a recombinant NS3 protein representative of ATPase/helicase domain from hepatitis C virus.
It has been shown that the Hepatitis C virus nonstructural NS3 protein possesses at least two enzymatic domains: a serine-protease domain and an adenosine triphosphatase (ATPase)/helicase domain. In this report, a truncated fragment of NS3 (26 kDa), representing main epitopes from the (ATPase)/helicase domain, has been expressed in Escherichia coli. The recombinant protein was purified by Ion Metal Affinity Chromatography (IMAC) with more than 90% purity. The recognition of B-cell linear epitopes in the NS3 protein was evaluated by immunoblot. The recombinant NS3 protein was reduced and carboxymethylated, and the recognition of either conformational and/or linear B-cell determinants was evaluated by ELISA. The inclusion of the recombinant NS3 protein in a third-generation diagnostic system UltraMicroELISA (UMELISA) allowed an increase in the sensitivity, due to the detection of a new variety of false-negative sera in blood donor test samples.
Pentón
N
N
Diagnostics Production Plant, Center for Genetic Engineering and Biotechnology, P.O. Box 6162, Havana 10600, Cuba.
Musacchio
A
A
Rivera
J M
JM
Roca
J
J
Ponce
M
M
Rodríguez
D
D
Caballero
A
A
Tallo
Y I
YI
Narciandi
R E
RE
D000251:Adenosine Triphosphatases / Q000737:chemistry*; D002846:Chromatography, Affinity; D003001:Cloning, Molecular; D004265:DNA Helicases / Q000737:chemistry*; D004591:Electrophoresis, Polyacrylamide Gel; D004797:Enzyme-Linked Immunosorbent Assay; D000939:Epitopes; D004926:Escherichia coli / Q000378:metabolism; D005285:Fermentation; D015151:Immunoblotting; D008957:Models, Genetic; D010957:Plasmids / Q000378:metabolism; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D013696:Temperature; D017361:Viral Nonstructural Proteins / Q000737:chemistry* / Q000235:genetics*; D012697:Serine Endopeptidases; D043583:Nucleoside-Triphosphatase; D053487:DEAD-box RNA Helicases; D000085242:Viral Proteases
D000251:Adenosine Triphosphatases; D004265:DNA Helicases; D000939:Epitopes; D011994:Recombinant Proteins; D017361:Viral Nonstructural Proteins; C084422:NS3 protein, hepatitis C virus; D012697:Serine Endopeptidases; D043583:Nucleoside-Triphosphatase; D053487:DEAD-box RNA Helicases; D000085242:Viral Proteases