Determination of the minimal length of preS1 epitope recognized by a monoclonal antibody which inhibits attachment of Hepatitis B virus to hepatocytes

Sominskaya, Irina; Pushko, Peter; Dreilina, Dzidra; Kozlovskaya, T.; Pumpen, P.

doi:10.1007/bf00215767

Cited by 54 publications

(45 citation statements)

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“…Individual envelope proteins were detected with the following antibodies: human monoclonal antibody 4/7B (31), recognizing an epitope around amino acids 178 to 186 of S protein (a gift of R. A. Heijtink, Organon Teknika, Oss, The Netherlands); mouse monoclonal antibody S26 (25), specific for an epitope around the motif QDPR in the pre-S2 region (provided by V. Bichko, Scriptgen, Waltham, Mass. ); and mouse monoclonal antibody MA18/07 (16,37), directed against an epitope close to the N terminus of the pre-S1 domain (a gift of W. Gerlich). Detection was performed with peroxidaseconjugated secondary antibodies and the ECL-Plus system.…”

Section: Methodsmentioning

confidence: 99%

Hepatitis B Virus (HBV) Virion and Covalently Closed Circular DNA Formation in Primary Tupaia Hepatocytes and Human Hepatoma Cell Lines upon HBV Genome Transduction with Replication-Defective Adenovirus Vectors

Ren

Nassal

2001

J Virol

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Hepatitis B virus (HBV), the causative agent of B-type hepatitis in humans, is a hepatotropic DNAcontaining virus that replicates via reverse transcription. Because of its narrow host range, there is as yet no practical small-animal system for HBV infection. The hosts of the few related animal viruses, including woodchuck hepatitis B virus and duck hepatitis B virus, are either difficult to keep or only distantly related to humans. Some evidence suggests that tree shrews (tupaias) may be susceptible to infection with human HBV, albeit with low efficiency. Infection efficiency depends on interactions of the virus with factors on the surface and inside the host cell. To bypass restrictions during the initial entry phase, we used recombinant replicationdefective adenovirus vectors, either with or without a green fluorescent protein marker gene, to deliver complete HBV genomes into primary tupaia hepatocytes. Here we show that these cells, like the human hepatoma cell lines HepG2 and Huh7, are efficiently transduced by the vectors and produce all HBV gene products required to generate the secretory antigens HBsAg and HBeAg, replication-competent nucleocapsids, and enveloped virions. We further demonstrate that covalently closed circular HBV DNA is formed. Therefore, primary tupaia hepatocytes support all steps of HBV replication following deposition of the genome in the nucleus, including the intracellular amplification cycle. These data provide a rational basis for in vivo experiments aimed at developing tupaias into a useful experimental animal system for HBV infection. Hepatitis B virus (HBV), an enveloped DNA-containing virus that replicates through an RNA intermediate (38), is the type member of the family Hepadnaviridae and the causative agent of B-type hepatitis in humans. Chronic infections are widespread and associated with a greatly increased risk for the development of liver cirrhosis and, eventually, primary liver carcinoma (5, 35). Several fundamental aspects of the HBV replication cycle have been elucidated, mainly by genetic techniques using transfection of cloned HBV DNA into suitable human hepatoma cell lines, such as Huh7 and HepG2 (27,28). These cells support virus production, but neither they nor other cell lines can be infected. A related and similarly central problem is the lack of a feasible small-animal model of human HBV infection.All hepadnaviruses have very narrow host ranges. Efficient infection by HBV is well documented for only humans and chimpanzees and, in cell culture, for primary hepatocytes from these hosts. The current generic animal models, the woodchuck-woodchuck hepatitis B virus (WHV) and the pekin duck-duck hepatitis B virus systems (28), are useful in many respects but suffer from two different limitations. Woodchucks are difficult to keep, and their use is restricted to a few appropriately equipped facilities; even then, most studies have used outbred, wild-caught animals (33). Ducks, by contrast, are convenient experimental animals (19) but very distantly related to humans...

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Section: Methodsmentioning

confidence: 99%

Hepatitis B Virus (HBV) Virion and Covalently Closed Circular DNA Formation in Primary Tupaia Hepatocytes and Human Hepatoma Cell Lines upon HBV Genome Transduction with Replication-Defective Adenovirus Vectors

Ren

Nassal

2001

J Virol

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“…For this purpose, a short HBV preS1 epitope 31-DPAFRA-36 (or DPAFR, or DPAF) necessary and sufficient [160] to be recognized by monoclonal antibody MA18/7 [161] has been used. The behavior of the DPAFR epitope was systematically compared after introduction into all preferred insertion sites of the HBc molecule at positions 2, 78, 144, and 183 [103], and into the MIR carrying deletions of different length [136].…”

Section: Special Applications Of the Hbc Particle As An Epitope Carriermentioning

confidence: 99%

HBV Core Particles as a Carrier for B Cell/T Cell Epitopes

Pumpens¹,

Grens²

2001

Intervirology

244

195

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In the middle 80s, recombinant hepatitis B virus cores (HBc) gave onset to icosahedral virus-like particles (VLPs) as a basic class of non-infectious carriers of foreign immunological epitopes. The recombinant HBc particles were used to display immunodominant epitopes of hepatitis B, C, and E virus, human rhinovirus, papillomavirus, hantavirus, and influenza virus, human and simian immunodeficiency virus, bovine and feline leukemia virus, foot-and-mouth disease virus, murine cytomegalovirus and poliovirus, and other virus proteins, as well as of some bacterial and protozoan protein epitopes. Practical applicability of the HBc particles as carriers was enabled by their ability to high level synthesis and correct self-assembly in heterologous expression systems. The interest in the HBc VLPs was reinforced by the resolution of their fine structure by electron cryomicroscopy and X-ray crystallography, which revealed an unusual α-helical organization of dimeric units of HBc shells, alternative packing into icosahedrons with T = 3 and T = 4 symmetry, and the existence of long protruding spikes. The tips of the latter seem to be the optimal targets for the display of foreign sequences up to 238 amino acid residues in length. Combination of numerous experimental data on epitope display with the precise structural information enables a knowledge-based design of diagnostic, and vaccine and gene therapy tools on the basis of the HBc particles.

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“…An adx/adw serotype-specific neutralizing antibody, F35.25, was generated against the residues 32-53 in the preS1 (Petit et al 1989) and an adr serotype specific antibody KR127 against the residues 37-45 (Maeng et al 2000). On the other hand, a monoclonal antibody MA 18/7 specific for an ay serotype targets the residues 31-34 in the preS1 (Sominskaya et al 1992). Another ayw-specific 5a-19-1 monoclonal antibody can specifically interact with the residues 36-43 (Budkowska et al 1995).…”

Section: Discussionmentioning

confidence: 99%

Pre‐structured motifs in the natively unstructured preS1 surface antigen of hepatitis B virus

et al. 2007

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The preS1 surface antigen of hepatitis B virus (HBV) is known to play an important role in the initial attachment of HBV to hepatocytes. We have characterized structural features of the full-length preS1 using heteronuclear NMR methods and discovered that this 119-residue protein is inherently unstructured without a unique tertiary structure under a nondenaturing condition. Yet, combination of various NMR parameters shows that the preS1 contains ''pre-structured'' domains broadly covering its functional domains. The most prominent domain is formed by residues 27-45 and overlaps with the putative hepatocyte-binding domain (HBD) encompassing residues 21-47, within which two welldefined pre-structured motifs, formed by Pro 32 -Ala 36 and Pro 41 -Phe 45 are found. Additional, somewhat less prominent, pre-structured motifs are also formed by residues 11-18, 22-25, 37-40, and 46-50. Overall results suggest that the preS1 is a natively unstructured protein (NUP) whose N-terminal 50 residues, populated with multiple pre-structured motifs, contribute critically to hepatocyte binding.Keywords: hepatitis B virus; preS1; NMR; natively unstructured protein; pre-structured motif Supplemental material: see www.proteinscience.org Hepatitis B virus (HBV) is a member of the hepadnaviridae family (Ganem and Varmus 1987;Chisari et al. 1989) that poses a significant health threat to millions of people worldwide, particularly in Africa, Asia, and certain parts of Europe (Fung and Lok 2004;Wright 2006). Whereas HBV infection itself may not appear so fatal as AIDS or SARS chronic hepatitis B infection often develops into more serious symptoms such as cirrhosis, liver failure, and hepatocelluar carcinoma (Blumberg et al. 1989). While successful administration of HBV vaccines led to a significant reduction of HBV-related casualties (Stephenne 1990;Mahoney et al. 1993;Jilg 1998;Poland and Jacobson 2004), emergence of escape mutants or nonresponders to current vaccines argues for attempts to develop vaccines with better efficacy. In addition, more specific anti-HBV therapeutics are needed in order to completely eradicate HBV infection, since nucleoside drugs currently in use for HBV-related symptoms are not HBV specific, often causing side effects or inducing drug resistance. Efforts to discover anit-HBVspecific agents based upon natural products or siRNAs have been met with only partial success (Saag 2006;Shin et al. 2006).Even though it is generally accepted that HBV infection begins with an attachment of HBV surface antigens to a HBV-specific hepatocyte receptor, followed by subsequent entry of viral DNA into hepatocyte (Neurath 3 These authors contributed equally to this work. Reprint requests to: Kyou-Hoon Han, Molecular Cancer Research Center, Division of Molecular Therapeutics, KRIBB, Daejeon 305-806, Korea; e-mail: khhan600@kribb.re.kr; fax: 82-42-860-4259.Abbreviations: HBV, hepatitis B virus; NMR, nuclear magnetic resonance; NUP, natively unstructured protein; CSI, chemical-shift index.Article published online ahead of pr...

show abstract

Determination of the minimal length of preS1 epitope recognized by a monoclonal antibody which inhibits attachment of Hepatitis B virus to hepatocytes

Cited by 54 publications

References 28 publications

Hepatitis B Virus (HBV) Virion and Covalently Closed Circular DNA Formation in Primary Tupaia Hepatocytes and Human Hepatoma Cell Lines upon HBV Genome Transduction with Replication-Defective Adenovirus Vectors

Hepatitis B Virus (HBV) Virion and Covalently Closed Circular DNA Formation in Primary Tupaia Hepatocytes and Human Hepatoma Cell Lines upon HBV Genome Transduction with Replication-Defective Adenovirus Vectors

HBV Core Particles as a Carrier for B Cell/T Cell Epitopes

Pre‐structured motifs in the natively unstructured preS1 surface antigen of hepatitis B virus

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