scholarly journals Structural differences between the Woodchuck hepatitis virus core protein in dimer and capsid states indicate entropic and conformational regulation of assembly

2019 ◽  
Author(s):  
Zhongchao Zhao ◽  
Joseph Che-Yen Wang ◽  
Giovanni Gonzalez-Gutierrez ◽  
Balasubramanian Venkatakrishnan ◽  
Adam Zlotnick
Keyword(s):  
Life Cycle ◽  
Hepatitis B Virus ◽  
Hepatitis B ◽  
Drug Development ◽  
Capsid Protein ◽  
Hepatitis Virus ◽  
Model System ◽  
Woodchuck Hepatitis Virus ◽  
High Fidelity ◽  
B Virus

AbstractHepadnaviruses are hepatotropic enveloped DNA viruses with an icosahedral capsid. Hepatitis B virus (HBV) causes chronic infection in an estimated 240 million people; Woodchuck Hepatitis virus (WHV), an HBV homologue, has been an important model system for drug development. The dimeric capsid protein (Cp) plays multiple functions during the viral life cycle and thus has become an important target for a new generation of antivirals. Purified HBV and WHV Cp spontaneously assemble into 120-dimer capsids. Though they have 65% identity, WHV Cp has error-prone assembly with stronger protein-protein association. We have taken advantage of the differences in assembly to investigate the basis of assembly regulation. We have determined the structures of the WHV capsid to 4.5 Å resolution by cryo-EM and the WHV Cp dimer to 2.9 Å resolution by crystallography and examined the biophysical properties of the dimer. We found, in dimer, the subdomain that makes protein-protein interactions is partially disordered and rotated 21° from its position in capsid. This subdomain is susceptible to proteolysis, consistent with local disorder. These data show there is an entropic cost for assembly that is compensated for by the energetic gain of burying hydrophobic interprotein contacts. We propose a series of stages in assembly that incorporate disorder-to-order transition and structural shifts. WHV assembly shows similar susceptibility to HBV antiviral molecules, suggesting that HBV assembly follows similar transitions and indicating WHV’s importance as a model system for drug development. We suggest that a cascade of structural changes may be a common mechanism for regulating high fidelity capsid assembly in HBV and other viruses.Author summaryCapsid assembly is a key step in the Hepatitis B virus (HBV) life cycle; it requires sophisticated regulation to ensure the production of capsids and viruses with high-fidelity. HBV capsids are constructed from 120 self-assembling capsid protein dimers (Cp). It was hypothesized that changes in Cp structure regulated the assembly reaction. Here we determined structures of dimer and capsid for Woodchuck Hepatitis virus (WHV), an HBV homologue. We observed that the component of the dimer involved in subunit-subunit interactions undergoes a disorder-order transition and changes structure concomitant with assembly. Meanwhile WHV Cp displays similar susceptibility to HBV antiviral. We propose that this structural transition entropically regulates assembly and may be a common theme in other viruses.

scholarly journals Antiviral Activities of Oral 1-O-Hexadecylpropanediol-3-Phosphoacyclovir and Acyclovir in Woodchucks with Chronic Woodchuck Hepatitis Virus Infection

2000 ◽  
Vol 44 (7) ◽  
pp. 1964-1969 ◽  
Author(s):  
Karl Y. Hostetler ◽  
James R. Beadle ◽  
William E. Hornbuckle ◽  
Christine A. Bellezza ◽  
Ilia A. Tochkov ◽  
...  
Keyword(s):  
Hepatitis B Virus ◽  
Virus Infection ◽  
Hepatitis B ◽  
Hepatitis Virus ◽  
Response To Therapy ◽  
Hydroxyl Group ◽  
Woodchuck Hepatitis Virus ◽  
Acyclovir Treatment ◽  
B Virus ◽  
Hepatitis Virus Infection

ABSTRACT Acyclovir triphosphate is a potent inhibitor of hepatitis B virus DNA polymerase, but acyclovir treatment provides no benefit in patients with hepatitis B virus infection. This is due in part to the fact that hepatitis B virus, unlike herpes simplex virus, does not code for a viral thymidine kinase which catalyzes the initial phosphorylation of acyclovir. We synthesized 1-O-octadecyl-sn-glycero-3-phospho (3-P)-acyclovir and found that it was highly active in reducing hepatitis B virus replication in 2.2.15 cells, while acyclovir was inactive. The greater antiviral activity of 1-O-octadecyl-sn-glycero-3-P-acyclovir appeared to be due to liver cell metabolism of the compound to acyclovir monophosphate (K. Y. Hostetler et al., Biochem. Pharmacol. 53:1815–1822, 1997). However, a closely related compound without a hydroxyl group at the sn-2 position of glycerol, 1-O-hexadecylpropanediol-3-P-acyclovir, was more active and selective in 2.2.15 cells in vitro. In this study, we treated woodchucks chronically infected with woodchuck hepatitis virus with increasing oral doses of 1-O-hexadecylpropanediol-3-P-acyclovir and assessed the response to therapy versus acyclovir or a placebo. At a dosage of 10 mg/kg of body weight twice a day, the test compound significantly inhibited viral replication in vivo, as indicated by a 95% reduction in serum woodchuck hepatitis virus DNA levels and by a 54% reduction in levels of woodchuck hepatitis virus replicative intermediates in the liver. Higher doses were somewhat less effective. In contrast, 20 mg of acyclovir/kg twice daily, a 5.3-fold-higher molar dosage, had no demonstrable activity against woodchuck hepatitis virus. Oral 1-O-hexadecylpropanediol-3-P-acyclovir appeared to be safe and effective in chronic woodchuck hepatitis virus infection.

scholarly journals Woodchuck Hepatitis Virus Contains a Tripartite Posttranscriptional Regulatory Element

1998 ◽  
Vol 72 (6) ◽  
pp. 5085-5092 ◽  
Author(s):  
John E. Donello ◽  
Jonathan E. Loeb ◽  
Thomas J. Hope
Keyword(s):  
Hepatitis B Virus ◽  
Hepatitis B ◽  
Hepatitis Virus ◽  
Regulatory Element ◽  
Deletion Analysis ◽  
Woodchuck Hepatitis Virus ◽  
B Virus ◽  
Cytoplasmic Accumulation ◽  
Posttranscriptional Regulatory Element

ABSTRACT The hepatitis B virus posttranscriptional regulatory element (HBVPRE) is a cis-acting RNA element that partially overlaps with enhancer I and is required for the cytoplasmic accumulation of HBV surface RNAs. We find that the closely related woodchuck hepatitis virus (WHV), which has been shown to lack a functional enhancer I, also contains a posttranscriptional regulatory element (WPRE). Deletion analysis suggests that the WPRE consists of three independent subelements. Comparison of the bipartite HBVPRE and tripartite WPRE activities reveals that the tripartite WPRE is two to three times more active than the bipartite HBVPRE. Mutation of a single WPRE subelement decreases WPRE activity to the level of the HBVPRE. Bipartite and tripartite chimeras of the WPRE and HBVPRE possess activities which suggest that elements containing three subelements are posttranscriptionally stronger than those containing two. These data demonstrate that the posttranscriptional regulatory element is conserved within the mammalian hepadnaviruses and that its strength is determined by the number of subelements within the RNA.

scholarly journals In vivo antiviral activity and pharmacokinetics of (-)-cis-5-fluoro-1-[2-(hydroxymethyl)-1,3-oxathiolan-5-yl]cytosine in woodchuck hepatitis virus-infected woodchucks.

1997 ◽  
Vol 41 (10) ◽  
pp. 2076-2082 ◽  
Author(s):  
J M Cullen ◽  
S L Smith ◽  
M G Davis ◽  
S E Dunn ◽  
C Botteron ◽  
...  
Keyword(s):  
Hepatitis B Virus ◽  
Treatment Group ◽  
Hepatitis B ◽  
Dna Polymerase ◽  
Hepatitis Virus ◽  
Woodchuck Hepatitis Virus ◽  
Normal Ranges ◽  
B Virus

The (-) enantiomer of cis-5-fluoro-1l-[2-(hydroxymethyl)-1,3-oxathiolan-5-yl]cytosine [(-)-FTC)], a substituted oxathiolane compound with anti-hepatitis B virus activity in vitro, was assessed for its efficacy in woodchucks with naturally acquired woodchuck hepatitis virus (WHV) infection. Pharmacokinetics and in vitro anabolism were also determined. (-)-FTC was anabolized to the 5'-triphosphate in a dose-related fashion, reaching a maximum concentration at about 24 h in cultured woodchuck hepatocytes. Following administration of a dose of 10 mg/kg of body weight intraperitoneally (i.p.), the clearance of (-)-FTC from plasma was monoexponential, the terminal half-life was 3.76 +/- 1.4 h, and the systemic clearance was 0.12 +/- 0.06 liters/h/kg. The antiviral efficacy of (-)-FTC in the woodchuck model was assessed by quantitation of serum WHV DNA levels and by WHV particle-associated DNA polymerase activity at two dosages, 30 and 20 mg/kg given i.p. twice daily (b.i.d.), respectively. The level of WHV DNA in serum was reduced 20- to 150-fold (average, 56-fold) in the 30-mg/kg-b.i.d. treatment group and 6- to 49-fold (average, 27-fold) in the 20-mg/kg-b.i.d. treatment group. Viral DNA polymerase levels diminished accordingly. One week after treatment was discontinued, WHV levels returned to pretreatment levels in both studies. These animals were biopsied before and following treatment with 30 mg of (-)-FTC per kg. Their livers were characterized by a mild increase in cytoplasmic lipid levels, but this change was not associated with altered liver enzyme levels. Serum chemistry and hematology results were within the normal ranges for all treated animals. We conclude that (-)-FTC is a potent antihepadnaviral agent and that it has no detectable toxic effects in woodchucks when given for up to 25 days. Further development of (-)-FTC as an anti-hepatitis B virus therapy for patients is warranted.

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