scholarly journals Formation of a Functional Hepatitis B Virus Replication Initiation Complex Involves a Major Structural Alteration in the RNA Template

1998 ◽  
Vol 18 (11) ◽  
pp. 6265-6272 ◽  
Author(s):  
Jürgen Beck ◽  
Michael Nassal
Keyword(s):  
Hepatitis B Virus ◽  
Hepatitis B ◽  
Replication Initiation ◽  
State Structure ◽  
Duck Hepatitis ◽  
P Protein ◽  
Encapsidation Signal ◽  
B Virus ◽  
Covalently Linked

ABSTRACT The DNA genome of a hepatitis B virus is generated by reverse transcription of the RNA pregenome. Replication initiation does not involve a nucleic acid primer; instead, the hepadnavirus P protein binds to the structured RNA encapsidation signal ɛ, from which it copies a short DNA primer that becomes covalently linked to the enzyme. Using in vitro-translated duck hepatitis B virus (DHBV) P protein, we probed the secondary structure of the protein-bound DHBV ɛ RNA (Dɛ) and observed a marked conformational change compared to free Dɛ RNA. Several initiation-competent mutant RNAs with a different free-state structure were similarly altered, whereas a binding-competent but initiation-deficient variant was not, indicating the importance of the rearrangement for replication initiation and suggesting a mechanistic coupling to encapsidation.

scholarly journals Reconstitution of a Functional Duck Hepatitis B Virus Replication Initiation Complex from Separate Reverse Transcriptase Domains Expressed in Escherichia coli

2001 ◽  
Vol 75 (16) ◽  
pp. 7410-7419 ◽  
Author(s):  
Jürgen Beck ◽  
Michael Nassal
Keyword(s):  
Escherichia Coli ◽  
Hepatitis B Virus ◽  
Hepatitis B ◽  
Reverse Transcriptase ◽  
Replication Initiation ◽  
In Vitro Translation ◽  
Duck Hepatitis B Virus ◽  
Duck Hepatitis ◽  
P Protein ◽  
B Virus

ABSTRACT Hepatitis B viruses replicate through reverse transcription of an RNA intermediate, the pregenomic RNA (pgRNA). Replication is initiated de novo and requires formation of a ribonucleoprotein complex comprising the viral reverse transcriptase (P protein), an RNA stem-loop structure (ɛ) on the pgRNA, and cellular proteins, including the heat shock protein Hsp90, the cochaperone p23, and additional, as yet unknown, factors. Functional complexes catalyze the synthesis of a short DNA primer that is templated by ɛ and covalently linked to the terminal protein (TP) domain of P protein. Currently, the only system for generating such complexes in the test tube is in vitro translation of duck hepatitis B virus (DHBV) P protein in rabbit reticulocyte lysate (RRL), which also provides the necessary factors. However, its limited translation capacity precludes a closer analysis of the complex. To overcome this restriction we sought to produce larger amounts of DHBV P protein by expression in Escherichia coli, followed by complex reconstitution in RRL. Because previous attempts to generate full-length P protein in bacteria have failed we investigated whether separate expression of the TP and reverse transcriptase-RNase H (RT-RH) domains would allow higher yields and whether these domains could trans complement each other. Indeed, TP and, after minor C-terminal modifications, also RT-RH could be expressed in substantial amounts, and when added to RRL, they were capable of ɛ-dependent DNA primer synthesis, demonstrating posttranslational activation. This reconstitution system should pave the way for a detailed understanding of the unique hepadnaviral replication initiation mechanism.

Effect of 3′-azido-3′-deoxythymidine on replication of duck hepatitis B virus in vivo and in vitro

1989 ◽  
Vol 29 (4) ◽  
pp. 244-248 ◽  
Author(s):  
Hideaki Haritani ◽  
Toshikazu Uchida ◽  
Yasunori Okuda ◽  
Toshio Shikata
Keyword(s):  
Hepatitis B Virus ◽  
Hepatitis B ◽  
Duck Hepatitis B Virus ◽  
Duck Hepatitis ◽  
B Virus

scholarly journals Anti-Hepatitis B Virus Activity of Esculetin from Microsorium fortunei In Vitro and In Vivo

Molecules ◽  
2019 ◽  
Vol 24 (19) ◽  
pp. 3475 ◽  
Author(s):  
Si-Xin Huang ◽  
Jun-Fei Mou ◽  
Qin Luo ◽  
Qing-Hu Mo ◽  
Xian-Li Zhou ◽  
...  
Keyword(s):  
Hepatitis B Virus ◽  
Hepatitis B ◽  
Hepatitis B Surface Antigen ◽  
Antiviral Drug ◽  
Liver Carcinoma ◽  
Dependent Manner ◽  
Duck Hepatitis ◽  
B Virus

Coumarins are widely present in a variety of plants and have a variety of pharmacological activities. In this study, we isolated a coumarin compound from Microsorium fortunei (Moore) Ching; the compound was identified as esculetin by hydrogen and carbon spectroscopy. Its anti-hepatitis B virus (HBV) activity was investigated in vitro and in vivo. In the human hepatocellular liver carcinoma 2.2.15 cell line (HepG2.2.15) transfected with HBV, esculetin effecting inhibited the expression of the HBV antigens and HBV DNA in vitro. Esculetin inhibited the expression of Hepatitis B virus X (HBx) protein in a dose-dependent manner. In the ducklings infected with duck hepatitis B virus (DHBV), the levels of DHBV DNA, duck hepatitis B surface antigen (DHBsAg), duck hepatitis B e-antigen (DHBeAg), alanine aminotransferase (ALT) and aspartate aminotransferase (AST) decreased significantly after esculetin treatment. Summing up the above, the results suggest that esculetin efficiently inhibits HBV replication both in vitro and in vivo, which provides an opportunity for further development of esculetin as antiviral drug.

Duck hepatitis B virus polymerase gene mutants associated with resistance to lamivudine have a decreased replication capacity in vitro and in vivo

2001 ◽  
Vol 34 (1) ◽  
pp. 114-122 ◽  
Author(s):  
Béatrice Seignères ◽  
Stéphanie Aguesse-Germon ◽  
Christian Pichoud ◽  
Isabelle Vuillermoz ◽  
Catherine Jamard ◽  
...  
Keyword(s):  
Hepatitis B Virus ◽  
Hepatitis B ◽  
Replication Capacity ◽  
Polymerase Gene ◽  
Duck Hepatitis B Virus ◽  
Duck Hepatitis ◽  
B Virus

scholarly journals Interaction between ganciclovir and foscarnet as inhibitors of duck hepatitis B virus replication in vitro.

1996 ◽  
Vol 40 (5) ◽  
pp. 1180-1185 ◽  
Author(s):  
G Civitico ◽  
T Shaw ◽  
S Locarnini
Keyword(s):  
Hepatitis B Virus ◽  
Hepatitis B ◽  
Hbv Dna ◽  
Duck Hepatitis ◽  
Chronic Hepatitis B Virus ◽  
B Virus ◽  
Dose Limiting Toxicity

Safe and effective treatments for chronic hepatitis B virus (HBV) infection have yet to be developed. Both ganciclovir (9-[1,3-dihydroxy-2-propoxymethyl]guanine) and foscarnet (trisodium phosphonoformate hexahydrate) are potent inhibitors of hepadnavirus replication when used individually in vitro and in vivo. However, the clinical usefulness of each drug is reduced by dose-limiting toxicity, especially during long-term monotherapy. Here we demonstrate additive inhibition of duck HBV DNA replication in cultures of primary duck hepatocytes congenitally infected with duck HBV by combinations of ganciclovir and foscarnet at low, clinically achievable concentrations. These results suggest that the effects of ganciclovir and foscarnet against HBV may be additive in vivo.

scholarly journals In Vitro Reconstitution of a Functional Duck Hepatitis B Virus Reverse Transcriptase: Posttranslational Activation by Hsp90

2000 ◽  
Vol 74 (24) ◽  
pp. 11447-11455 ◽  
Author(s):  
Jianming Hu ◽  
Dana Anselmo
Keyword(s):  
Hepatitis B Virus ◽  
Hepatitis B ◽  
Reverse Transcriptase ◽  
Host Factors ◽  
Duck Hepatitis B Virus ◽  
Duck Hepatitis ◽  
B Virus ◽  
Protein Priming ◽  
Hsp90 Function

ABSTRACT Reverse transcription in hepatitis B viruses is initiated through a unique protein priming mechanism whereby the viral reverse transcriptase (RT) first assembles into a ribonucleoprotein (RNP) complex with its RNA template and then initiates DNA synthesis de novo using the RT itself as a protein primer. RNP formation and protein priming require the assistance of host cell factors, including the molecular chaperone heat shock protein 90 (Hsp90). To better understand the mechanism of RT activation by Hsp90, we have now mapped the minimal RT sequences of the duck hepatitis B virus that are required for chaperone binding, RNP formation, and protein priming. Furthermore, we have reconstituted in vitro both RNP formation and protein priming using purified RT proteins and host factors. Our results show that (i) Hsp90 recognizes two independent domains of the RT, both of which are necessary for RNP formation and protein priming; (ii) Hsp90 function is required not only to establish, but also to maintain, the RT in a state competent for RNA binding; and (iii) Hsp90 is not required during RT synthesis and can activate the RT posttranslationally. Based on these findings, we propose a model for Hsp90 function whereby the chaperone acts as an active interdomain bridge to bring the two RT domains into a poised but labile conformation competent for RNP formation. It is anticipated that the reconstitution system established here will facilitate the isolation of additional host factors required for RT functions and further elucidation of the mechanisms of RT activation.

The pregenome/C RNA of duck hepatitis B virus is not used for translation of core protein during the early phase of infection in vitro

2015 ◽  
Vol 196 ◽  
pp. 13-19 ◽  
Author(s):  
Qiang Liu ◽  
Juan Huang ◽  
Renyong Jia ◽  
Mingshu Wang ◽  
Dekang Zhu ◽  
...  
Keyword(s):  
Hepatitis B Virus ◽  
Hepatitis B ◽  
Early Phase ◽  
Core Protein ◽  
Duck Hepatitis B Virus ◽  
Duck Hepatitis ◽  
B Virus
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