• Lanford, Robert E (PI)

    Project: Research project

    Project Details


    Hepatitis B virus (HBV) represents a serious worldwide health problem. The
    acute infection is usually self-limiting but may prove fatal, and, in
    addition, up to 10% of infected individuals become chronic carriers of the
    disease. The total number of carriers has been estimated at 280 million.
    Chronicity frequently progresses to cirrhosis the liver and hepatocellular
    carcinoma. Chronic carriers represent the main reservoir of the virus, with
    transmission occurring through contaminated blood products, IV drug abuse,
    sexual contact, and from mother to infant at birth. A precise understanding
    of the mechanism of viral replication is required to devise strategies for
    eliminat-ing the carrier state. This proposal will address several
    unresolved issues pertaining to the replication of the viral genome and
    viral morphogenesis. The first specific aim is to examine core protein
    mutants for assembly and for encapsidation of pregenomic RNA, polymerase,
    and polymerase-RNA complexes. The proteins will be expressed in insect
    cells using a baculovirus expression system or in in vitro translation
    systems and in vitro assays will be developed to examine these functions.
    The ability of core mutants to complement the replication of a core minus
    HBV mutant will be examined in the human hepatocellular carcinoma cell
    line, HepG2. The mechanism of encapsidation and the functional significance
    of the virion associated kinase will be explored. The second specific aim
    will be to express polymerase and polymerase functional domains in insect
    cells and to develop in vitro assays for reverse transcription, pregenomic
    RNA binding, nucleotide binding, and RNase H activity. The third specific
    aim will be to examine the RNA sequence requirement for encapsidation of
    pregenomic RNA by core and for interaction of pregenomic RNA with
    polymerase. The in vitro RNA transcripts used in assays for core and
    polymerase functions will be altered to determine the minimal sequences
    required for recognition. The final specific aim will be to utilize the
    information obtained in the in vitro assays to make specific mutations in
    the HBV genome and examine replication the mutants in HepG2 cells.
    Effective start/end date12/5/901/31/02


    • National Institutes of Health: $218,994.00
    • National Institutes of Health: $310,695.00
    • National Institutes of Health: $335,909.00
    • National Institutes of Health: $231,870.00
    • National Institutes of Health: $287,253.00


    • Medicine(all)


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