Secretion and purification of hepatitis C virus NS1 glycoprotein produced by recombinant baculovirus-infected insect cells

Gene. 1993 Jul 30;129(2):207-14. doi: 10.1016/0378-1119(93)90270-d.

Abstract

Recombinant baculoviruses that produce a putative non-structural protein 1 (NS1) of hepatitis C virus (HCV), predicted to be the second envelope glycoprotein, were constructed. The recombinant NS1 protein (re-NS1) produced in infected insect cells was localized on the cell surface and was apparently glycosylated, because it was susceptible to treatment with both tunicamycin and N-glycanase. Furthermore, re-NS1 was effectively secreted into the culture supernatant when the putative NS1 signal peptide (SP) was replaced by the SP of rabies virus G protein, and the C-terminal hydrophobic region was eliminated. The secreted re-NS1 was tagged with six His residues at the C terminus and purified simply by native Ni(2+)-nitrilotriacetic acid (Ni(2+)-NTA) affinity column chromatography. An enzyme-linked immunosorbent assay (ELISA) was developed for the serological diagnosis of HC using purified re-NS1. Anti-NS1 antibody (Ab) was detected in 55 of 60 patients (92%) with chronic HC liver diseases. Thus, this ELISA for Ab directed against HCV re-NS1 produced in insect cells is useful for the detection of chronic HC patients.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Animals
  • Antigens, Viral / biosynthesis*
  • Baculoviridae
  • Base Sequence
  • Chromatography, Affinity / methods
  • DNA, Recombinant
  • Enzyme-Linked Immunosorbent Assay
  • Glycoproteins / biosynthesis
  • Hepacivirus / chemistry*
  • Hepatitis Antibodies / immunology
  • Hepatitis C / diagnosis
  • Hepatitis C / immunology
  • Hepatitis C Antibodies
  • Hepatitis C Antigens
  • Humans
  • Insecta
  • Molecular Sequence Data
  • Protein Sorting Signals
  • Recombinant Fusion Proteins / biosynthesis
  • Viral Envelope Proteins / biosynthesis
  • Viral Nonstructural Proteins / biosynthesis*
  • Viral Nonstructural Proteins / genetics
  • Viral Nonstructural Proteins / isolation & purification

Substances

  • Antigens, Viral
  • DNA, Recombinant
  • Glycoproteins
  • Hepatitis Antibodies
  • Hepatitis C Antibodies
  • Hepatitis C Antigens
  • Protein Sorting Signals
  • Recombinant Fusion Proteins
  • Viral Envelope Proteins
  • Viral Nonstructural Proteins
  • glycoprotein G, Rabies virus

Associated data

  • GENBANK/D13043
  • GENBANK/D13406
  • GENBANK/L03653
  • GENBANK/L03654
  • GENBANK/L05530
  • GENBANK/L07632
  • GENBANK/M96931
  • GENBANK/M96932
  • GENBANK/M96933
  • GENBANK/X54209