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Query: UMLS:C0019163 (hepatitis B)
 38,309 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

A recombinant baculovirus expression vector was constructed to express the core (capsid) protein of the hepatitis B virus. Along with the expected 21-kDa polypeptide, a second 24-kDa protein was observed. Immunoprecipitation and immunoblotting using a rabbit polyclonal anticore antiserum demonstrated that the two proteins were related. The core gene originally was cloned in-frame with the polyhedrin initiator codon that had been altered to AUU as a means of preventing fusion protein formation. A transient expression assay revealed expression of the 24-kDa protein was prevented if a frame-shift mutation was created upstream of the HBV core translation start site. These results suggest that the 24-kDa protein was the result of an unexpectedly high level of translation initiation at the AUU codon that gave rise to a polyhedrin-HBV core fusion protein. The 24-kDa core protein was shown to be a polyhedrin fusion protein by immunoblotting with an antipolyhedrin antiserum, and initiation at the AUU was demonstrated by amino terminal protein sequencing. Methods to prevent undesired fusion protein expression using this or similar vectors are discussed.
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PMID:Polyhedrin initiator codon altered to AUU yields unexpected fusion protein from a baculovirus vector. 193 Oct 38

We constructed a yeast strain that simultaneously expresses four genes encoding the major S, middle S, large S hepatitis B viral envelope proteins and the core protein under the control of the yeast glyceraldehyde-3-phosphate dehydrogenase promoter and terminator. The lysate from this cell line, examined by immunological, physicochemical methods and electron microscopy, was found to contain spherical particles with a diameter of about 40 nm and a density of 1.25 g/ml. These particles reacted with anti-envelope antibodies, but not with anti-core antibodies. However, core antigenicity appeared upon treatment with 3% Nonidet P-40 that eliminates an outer envelope. These observations suggest production of a virion-like complex structure, or at least its DNA-less analog, consisting of core particle enveloped by antibody-reactive envelope. Such a structure was made only when all the four gene products were synthesized in a yeast cell. This system may be useful for the study of virus structure and assembly, and for improved vaccine development.
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PMID:Production of hepatitis B virion-like particles in yeast. 193 46

Human hepatitis B virus encodes a secretory core protein, referred to as the HBe protein, whose secretion is mediated by the pre-C signal sequence. Here we examined whether this sequence is important only for translocation of the HBe precursor (the precore protein) or whether it also contributes to the structural and biophysical properties of the mature HBe protein. When a truncated hepatitis B virus precore protein, lacking the basic C-terminal domain which is cleaved from the wild-type protein during its conversion into HBe, was expressed in human hepatoma cells, only a small amount of HBe-like protein was produced. This protein was slightly smaller than the wild-type HBe protein, suggesting that C-terminal cleavage of the precore protein does not occur at the suggested site. When the authentic signal sequence of the precore protein (the pre-C sequence) was replaced by the unrelated signal sequence of an influenza virus hemagglutinin, not only the full-length but also the C-terminally truncated protein was expressed and secreted with high efficiency. Western blot (immunoblot) analyses with nonreducing gels and conformation-specific monoclonal antibodies revealed that the HBe protein secreted under control of the pre-C signal sequence was a monomer with HBe antigenicity, whereas the HBe-like protein secreted under control of the hemagglutinin signal sequence was a disulfide-bridge-linked dimer with both HBe and HBc antigenicity. Electron microscopic examination of gradient-purified particulate core gene products showed that HBe protein secreted under control of the hemagglutinin signal sequence forms core particles, whereas HBe protein secreted under control of the pre-C sequence does not. Thus, the pre-C sequence not only mediates the secretion but also determines the structural and aggregational properties of the HBe protein.
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PMID:The quaternary structure, antigenicity, and aggregational behavior of the secretory core protein of human hepatitis B virus are determined by its signal sequence. 194 54

The biosynthesis of the secretory core gene product of the duck hepatitis B virus (DHBe protein) was examined. Recombinant vaccinia viruses were constructed encoding either the full-length or C-terminally truncated forms of the DHBe precursor protein (precore protein) and used to express these proteins in the human hepatoma cell line HepG2. Western immunoblot analysis of core gene products isolated from cells producing the full-length precore protein revealed the presence of DHBe precursor proteins containing the strongly basic C-terminal sequence which is lacking in the mature DHBe protein. These proteins were not secreted, suggesting that C-terminal proteolytic processing of the precore protein represents an obligatory step for DHBe biosynthesis. Pulse-chase experiments showed that this cleavage reaction occurs late during DHBe synthesis. Interestingly, when mutated precore proteins were expressed which lacked the basic C-terminal domain, proteins were produced which were glycosylated but not secreted. This shows that the transient presence of this region is essential for intracellular transport of the precore protein. Cell sorter analyses revealed that production of a cell surface-expressed variant of the secretory core protein is a feature conserved between the duck and the human hepatitis B viruses. Surprisingly, the C terminus of the membrane-expressed DHBe protein was accessible from the outside, showing that the topology of this interesting protein is more complicated than expected.
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PMID:Biosynthesis of the secretory core protein of duck hepatitis B virus: intracellular transport, proteolytic processing, and membrane expression of the precore protein. 204 Oct 77

As a step toward understanding the assembly of the hepatitis B virus (HBV) nucleocapsid at a molecular level, we sought to define the primary sequence requirements for assembly of the HBV core protein. This protein can self assemble upon expression in Escherichia coli. Applying this system to a series of C-terminally truncated core protein variants, we mapped the C-terminal limit for assembly to the region between amino acid residues 139 and 144. The size of this domain agrees well with the minimum length of RNA virus capsid proteins that fold into an eight-stranded beta-barrel structure. The entire Arg-rich C-terminal domain of the HBV core protein is not necessary for assembly. However, the nucleic acid content of particles formed by assembly-competent core protein variants correlates with the presence or absence of this region, as does particle stability. The nucleic acid found in the particles is RNA, between about 100 to some 3,000 nucleotides in length. In particles formed by the full-length protein, the core protein mRNA appears to be enriched over other, cellular RNAs. These data indicate that protein-protein interactions provided by the core protein domain from the N terminus to the region around amino acid 144 are the major factor in HBV capsid assembly, which proceeds without the need for substantial amounts of nucleic acid. The presence of the basic C terminus, however, greatly enhances encapsidation of nucleic acid and appears to make an important contribution to capsid stability via protein-nucleic acid interactions. The observation of low but detectable levels of nucleic acid in particles formed by core protein variants lacking the Arg-rich C terminus suggests the presence of a second nucleic acid-binding motif in the first 144 amino acids of the core protein. Based on these findings, the potential importance of the C-terminal core protein region during assembly in vivo into authentic, replication-competent nucleocapsids is discussed.
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PMID:Hepatitis B virus nucleocapsid assembly: primary structure requirements in the core protein. 219 Nov 49

Permanent mouse fibroblast LTK- cells were transfected with dimeric hepatitis B virus (HBV) DNA linked to the simian virus 40 (SV40) early promoter/enhancer. Many clones stably expressed high levels of polyadenylated RNAs encoding hepatitis B surface (HBs) proteins (2.1 kb), HBe protein (3.6 kb), and HBx protein (0.6 kb). Although a chimeric RNA (4.0 kb) probably starting from the SV40 promoter was also synthesized, transcription of viral RNAs was predominantly directed by HBV promoters and its terminator. In contrast to HBV-transfected liver cells, the fibroblasts expressed only pregenomic 3.6-kb transcripts starting 5' to, but not within, the precore sequence. Thus, no normal core protein could be synthesized, but the cells expressed and secreted HBe protein of heterogeneous size. Small and middle HBs proteins were strongly expressed, while large HBs protein was almost absent. HBx mRNA expression was more efficient in mouse fibroblasts than in human hepatoma cells and 18-kDa HBx protein was exclusively detected in purified nuclei. Expression of HBe, small and middle HBs, and HBx proteins apparently does not require hepatic factors. Underexpression of HBc mRNA and large HBs mRNA suggests that activity of their promoters depends on cell-type-specific transcription factors.
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PMID:Expression pattern of the hepatitis B virus genome in transfected mouse fibroblasts. 221 24

Permanent murine fibroblasts (LTK-) were transfected with a dimer of hepatitis B virus (HBV) DNA and a neomycin resistance gene which were both linked to the simian virus 40 (SV40) early promoter/enhancer. One of the stably transfected clones, LTK4/36, which secreted HBsAg, HBeAg, and HBV DNA was further analyzed. It contained eight to nine copies of integrated HBV DNA per haploid genome and low amounts of episomal HBV DNA. The secreted viral DNA was covalently linked to protein and was associated with particles which had the characteristic density of natural virions from serum of human viremic carriers. The particles contained an endogenous DNA polymerase, small and middle surface proteins, but in contrast to natural virions very little core protein and large surface protein. Instead of core protein, they contained incompletely processed HBe protein which is colinear to core protein. The fibroblast-derived virions were less stable than virions from human carriers or from transfected hepatoma cells. After several days of storage, their DNA was only partially protected against DNase. Obviously, nonhepatic cells can express HBV-like particles, even if liver-dependent gene products like large surface protein and core protein are missing.
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PMID:Replication of hepatitis B virus in transfected nonhepatic cells. 221 25

Hepatitis B virus DNA replicates via its own polymerase that also acts as reverse transcriptase (Summers and Mason, 1982). This enzyme is encoded by a 3.5 Kb mRNA transcript covering the whole genome. Since the same transcript also codes for the core protein, and since the core open reading frame (ORF) is located upstream of the pol ORF, it has been suggested that the polymerase is first produced as a core-pol fusion protein that subsequently undergoes cleavage. This is already known to be the case with retrovirus reverse transcriptase, for which a gag-pol fusion protein is made first and the latter protein is liberated by proteolytic cleavage. We investigated this problem using mutants that were modified at the translation initiation codon for the core and precore ORF. Our findings suggested that polymerase translation occurred from the internal AUG codon independently of core protein synthesis, and that obligatory production of the core-pol fusion protein is accordingly unlikely.
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PMID:Translation of hepatitis B virus DNA polymerase from the internal AUG codon, not from the upstream AUG codon for the core protein. 222 32

Precore and core proteins are two related co-carboxy-terminal proteins of hepatitis B virus. Precore protein contains the entire sequence of core protein plus an amino-terminal extension of 29 amino acid residues. Both proteins can display a common antigenic determinant known as core antigen (HBcAg). Clinically, HBcAg is detected in the nucleus, cytoplasm, or both of hepatitis B virus-infected hepatocytes. In order to understand the mechanism that regulates nuclear transport of HBcAg, various portions of precore and core proteins were linked to a reporter protein, human alpha-globin, and expressed in mammalian cells. Our results indicate that the precore protein-specific sequence, although important for nuclear transport, does not contain a nuclear localization signal. Instead, a signal for nuclear transport is located near the carboxy termini of precore and core proteins in the arginine-rich domain. This signal is made up of a set of two direct PRRRRSQS repeats and is highly conserved among mammalian hepadnaviruses.
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PMID:The arginine-rich domain of hepatitis B virus precore and core proteins contains a signal for nuclear transport. 224 90

Hepatitis B virus (HVB) is the prototype member of the hepadnaviridae, a family of small enveloped DNA viruses that replicate by reverse transcription. Assembly of replication-competent HBV nucleocapsids is based on specific interactions between the core protein, the product(s) of the P gene, and the RNA pregenome, which is marked for encapsidation by containing a sequence near its 5' end that acts in cis as an encapsidation signal. However, HBV produces several additional, almost identical, genomic transcripts that also bear the encapsidation sequence, but that are not encapsidated. The mechanism underlying this selection process has remained mysterious. Here we demonstrate that translating 80S ribosomes (but not scanning 40S ribosomal subunits) advancing into the encapsidation signal prevent its functioning. This finding reveals translational modulation of RNA function as a further regulatory mechanism employed by hepadnaviruses to utilize efficiently the restricted coding capacity of their extremely compact genome.
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PMID:Translational inactivation of RNA function: discrimination against a subset of genomic transcripts during HBV nucleocapsid assembly. 226 46


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