Fraction-specific populations of the hypervariable region of the hepatitis C virus in a patient with cryoglobulinemia.
Nucleotide sequences of the hypervariable region (HVR) of the E2/NS1 gene of hepatitis C virus (HCV), which are now thought to contain epitopes for neutralizing antibodies, were compared between antibody-bound HCV and free HCV in a patient with type II cryoglobulinemia. Antibody-bound HCV was immunoprecipitated with anti-human immunoglobulins from serum of the patient. Total RNA was recovered from the pellet and the supernatant, respectively, and the envelope gene containing the HVR was amplified by the reverse transcription and nested polymerase chain reaction. The amplified cDNA was examined by the single strand conformation polymorphism (SSCP) analysis. Sequences of bands separated by SSCP analysis were determined by the dideoxy chain termination method. SSCP analyses revealed that the HCV populations were completely different between antibody-bound HCV and free HCV: antibody-bound HCV was composed of two bands and free HCV was composed of three bands. These five bands showed different mobility with each other on the SSCP gel. Sequencing of each band revealed distinct HVR sequences, differing in 1-34 nucleotides and 1-15 deduced amino acids. Three sequences of free HCV was similar with each other (1-5 nucleotide and 1-4 amino acid differences). On the other hand, two sequences of antibody-bound HCV had 5-34 nucleotide and 5-15 amino acid differences with free HCV. Thirteen amino acids in the 5' of HVR were completely identical in three sequences of free HCV, whereas there were three and seven amino acid differences in two sequences of antibody-bound HCV. These findings suggest that isolated specific epitopes for envelope antibodies exist within the HVR.(ABSTRACT TRUNCATED AT 250 WORDS)
Kurosaki
M
M
Second Department of Internal Medicine, Faculty of Medicine, Tokyo Medical and Dental University, Japan.
Enomoto
N
N
Nouchi
T
T
Sakuma
I
I
Marumo
F
F
Sato
C
C
D000595:Amino Acid Sequence; D001483:Base Sequence; D003449:Cryoglobulinemia / Q000150:complications* / Q000821:virology; D017931:DNA Primers; D004279:DNA, Viral / Q000235:genetics; D016174:Hepacivirus / Q000235:genetics* / Q000302:isolation & purification; D006526:Hepatitis C / Q000150:complications / Q000821:virology*; D006801:Humans; D008969:Molecular Sequence Data; D010802:Phylogeny; D018807:Polymorphism, Single-Stranded Conformational; D014158:Transcription, Genetic; D014759:Viral Envelope Proteins / Q000235:genetics
D017931:DNA Primers; D004279:DNA, Viral; D014759:Viral Envelope Proteins; C091015:glycoprotein E2, Hepatitis C virus