Characterization of the genome and structural proteins of hepatitis C virus resolved from infected human liver

• Published in: Journal of general virology 2004-06, Vol.85 (6), p.1497-1507
• Main Authors: Nielsen, Soren U, Bassendine, Margaret F, Burt, Alastair D, Bevitt, Debra J, Toms, Geoffrey L
• Format: Article
• Language: English
• Subjects: Base Sequence; Biological and medical sciences; Dimerization; Fundamental and applied biological sciences. Psychology; Genome, Viral; Glycosylation; Hepacivirus - chemistry; Hepacivirus - genetics; Hepatitis C - virology; Hepatitis C virus; Human viral diseases; Humans; Infectious diseases; Liver - virology; Medical sciences; Microbiology; Miscellaneous; Molecular Sequence Data; RNA, Viral - analysis; Viral diseases; Viral Envelope Proteins - analysis; Viral hepatitis; Viral Structural Proteins - analysis; Viral Structural Proteins - chemistry; Virology
• ISSN: 0022-1317; 1465-2099
• DOI: 10.1099/vir.0.79967-0

1 School of Clinical Medical Sciences, Medical School, Framlington Place, Newcastle upon Tyne NE2 4HH, UK 2 School of Clinical and Laboratory Sciences, Medical School, Framlington Place, Newcastle upon Tyne NE2 4HH, UK Correspondence Søren U. Nielsen s.u.nielsen{at}newcastle.ac.uk In the absence of satisfactory cell culture systems for hepatitis C virus (HCV), virtually all that is known about the proteins of the virus has been learned by the study of recombinant proteins. Characterization of virus proteins from patients with HCV has been retarded by the low virus titre in blood and limited availability of infected tissue. Here, the authors have identified a primary infection in a liver transplanted into an immunodeficient patient with chronic HCV. The patient required re-transplant and the infected liver, removed 6 weeks after the initial transplant, had a very high titre of HCV, 5 x 10 9 International Units (IU) per gram of liver. The density distribution of HCV in iodixanol gradients showed a peak at 1·04 g ml –1 with 73 % of virus below 1·08 g ml –1 . Full-length HCV RNA was detected by Northern blotting and the ratio between positive- and negative-strand HCV RNA was determined as 60. HCV was partially purified by precipitation with heparin/Mn 2+ and a single species of each of the three structural proteins, core, E1 and E2, was detected by Western blotting. The molecular mass of core was 20 kDa, which corresponds to the mature form from recombinant sources. The molecular mass of glycoprotein E1 was 31 kDa before and 21 kDa after deglycosylation with PNGase F or endoglycosidase H. Glycoprotein E2 was 62 kDa before and 36 kDa after deglycosylation, but E2-P7 was not detected. This was in contrast to recombinant sources of E2 which contain E2-P7. EMBL accession numbers for sequences of HCV from infected human livers S6a and S6b are AJ557443 and AJ557444, respectively. Present address: Department of Rheumatology, Medical School, University of Newcastle, Newcastle upon Tyne, UK.