Biomaterial Database

Generation of infectious nucleocapsids by in vitro assembly of the shell protein on to the polymerase complex of the dsRNA bacteriophage phi 6. 1991

V M Olkkonen, and P M Ojala, and D H Bamford

Department of Genetics, University of Helsinki, Finland.

A method for the in vitro uncoating of the phi 6 nucleocapsid (NC) was developed. The resulting particle, designated as the NC core, containing the genomic double-stranded (ds) RNA segments and the proteins P1, P2, P4 and P7, was not infectious but had a highly enhanced in vitro transcriptase activity compared to that of the intact NC. The NC shell protein P8 was purified by immunoaffinity chromatography, and it was shown to self-assemble to shell-like structures upon addition of calcium ions. The conditions for the self-assembly of the shell were optimized. Shell reassembly on to the NC cores restored the infectivity but resulted in a decrease of transcriptase activity. No reassembly of the shell on to RNA-less cores (procapsids) produced from a cDNA construction in Escherichia coli was observed. Our results suggest that the intracellular uncoating of the NC is the event activating the phi 6 dsRNA transcriptase and that the NC shell is necessary for infectivity, probably for the passage of the NC through the host cytoplasmic membrane. Packaging of the dsRNA segments into the procapsid appears to be a prerequisite for NC shell assembly.

UI	MeSH Term	Description	Entries
D011189	Potassium Chloride	A white crystal or crystalline powder used in BUFFERS; FERTILIZERS; and EXPLOSIVES. It can be used to replenish ELECTROLYTES and restore WATER-ELECTROLYTE BALANCE in treating HYPOKALEMIA.	Slow-K,Chloride, Potassium
D002122	Calcium Chloride	A salt used to replenish calcium levels, as an acid-producing diuretic, and as an antidote for magnesium poisoning.	Calcium Chloride Dihydrate,Calcium Chloride, Anhydrous
D002213	Capsid	The outer protein protective shell of a virus, which protects the viral nucleic acid. Capsids are composed of repeating units (capsomers or capsomeres) of CAPSID PROTEINS which when assembled together form either an icosahedral or helical shape.	Procapsid,Prohead,Capsids,Procapsids,Proheads
D002846	Chromatography, Affinity	A chromatographic technique that utilizes the ability of biological molecules, often ANTIBODIES, to bind to certain ligands specifically and reversibly. It is used in protein biochemistry. (McGraw-Hill Dictionary of Scientific and Technical Terms, 4th ed)	Chromatography, Bioaffinity,Immunochromatography,Affinity Chromatography,Bioaffinity Chromatography
D004591	Electrophoresis, Polyacrylamide Gel	Electrophoresis in which a polyacrylamide gel is used as the diffusion medium.	Polyacrylamide Gel Electrophoresis,SDS-PAGE,Sodium Dodecyl Sulfate-PAGE,Gel Electrophoresis, Polyacrylamide,SDS PAGE,Sodium Dodecyl Sulfate PAGE,Sodium Dodecyl Sulfate-PAGEs
D006863	Hydrogen-Ion Concentration	The normality of a solution with respect to HYDROGEN ions; H+. It is related to acidity measurements in most cases by pH	pH,Concentration, Hydrogen-Ion,Concentrations, Hydrogen-Ion,Hydrogen Ion Concentration,Hydrogen-Ion Concentrations
D001435	Bacteriophages	Viruses whose hosts are bacterial cells.	Phages,Bacteriophage,Phage
D012321	DNA-Directed RNA Polymerases	Enzymes that catalyze DNA template-directed extension of the 3'-end of an RNA strand one nucleotide at a time. They can initiate a chain de novo. In eukaryotes, three forms of the enzyme have been distinguished on the basis of sensitivity to alpha-amanitin, and the type of RNA synthesized. (From Enzyme Nomenclature, 1992).	DNA-Dependent RNA Polymerases,RNA Polymerases,Transcriptases,DNA-Directed RNA Polymerase,RNA Polymerase,Transcriptase,DNA Dependent RNA Polymerases,DNA Directed RNA Polymerase,DNA Directed RNA Polymerases,Polymerase, DNA-Directed RNA,Polymerase, RNA,Polymerases, DNA-Dependent RNA,Polymerases, DNA-Directed RNA,Polymerases, RNA,RNA Polymerase, DNA-Directed,RNA Polymerases, DNA-Dependent,RNA Polymerases, DNA-Directed
D012330	RNA, Double-Stranded	RNA consisting of two strands as opposed to the more prevalent single-stranded RNA. Most of the double-stranded segments are formed from transcription of DNA by intramolecular base-pairing of inverted complementary sequences separated by a single-stranded loop. Some double-stranded segments of RNA are normal in all organisms.	Double-Stranded RNA,Double Stranded RNA,RNA, Double Stranded
D012367	RNA, Viral	Ribonucleic acid that makes up the genetic material of viruses.	Viral RNA