A micromolar pool of antigenically distinct precursors is required to initiate cooperative assembly of hepatitis B virus capsids in Xenopus oocytes.
Assembly of hepatitis B virus capsid-like (core) particles occurs efficiently in a variety of heterologous systems via aggregation of approximately 180 molecules of a single 21.5-kDa core protein (p21.5), resulting in an icosahedral capsid structure with T = 3 symmetry. Recent studies on the assembly of hepatitis B virus core particles in Xenopus oocytes suggested that dimers of p21.5 represent the major building block from which capsids are generated. Here we determined the concentration dependence of this assembly process. By injecting serially diluted synthetic p21.5 mRNA into Xenopus oocytes, we expressed different levels of intracellular p21.5 and monitored the production of p21.5 dimers and capsids by radiolabeling and immunoprecipitation, by radioimmunoassay, or by quantitative enzyme-linked immunosorbent assay analysis. The data revealed that (i) p21.5 dimers and capsids are antigenically distinct, (ii) capsid assembly is a highly cooperative and concentration-dependent process, and (iii) p21.5 must accumulate to a signature concentration of approximately 0.7 to 0.8 microM before capsid assembly initiates. This assembly process is strikingly similar to the assembly of RNA bacteriophage R17 as defined by in vitro studies.
Seifer
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Hormone Research Institute, University of California, San Francisco 94143-0534.
Zhou
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S
Standring
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D000818:Animals; D000956:Antigens, Viral / Q000276:immunology / Q000378:metabolism*; D002213:Capsid / Q000096:biosynthesis* / Q000276:immunology; D004305:Dose-Response Relationship, Drug; D006515:Hepatitis B virus / Q000254:growth & development*; D046911:Macromolecular Substances; D008845:Microinjections; D009865:Oocytes / Q000382:microbiology; D011487:Protein Conformation; D011498:Protein Precursors / Q000378:metabolism*; D012333:RNA, Messenger / Q000378:metabolism / Q000494:pharmacology; D012015:Reference Standards; D014758:Viral Core Proteins / Q000096:biosynthesis* / Q000276:immunology; D014981:Xenopus
D000956:Antigens, Viral; D046911:Macromolecular Substances; D011498:Protein Precursors; D012333:RNA, Messenger; D014758:Viral Core Proteins; C069130:nucleocapsid protein, Hepatitis C virus