Isolation and properties of highly purified Halobacterium cutirubrum deoxyribonucleic acid-dependent ribonucleic acid polymerase.
1. The subunits alpha and beta of Halobacterium cutirubrum DNA-dependent RNA polymerase have been purified to electrophoretic homogeneity. Both have mol.wt. 18000 and they are required in equimolar amounts for optimum activity. 2. The instability of the complete enzyme, alphabeta, in the absence of salt is due to the rapid inactivation of the beta subunit in these conditions. 3. Nearest-neighbour analysis of the product formed on poly[d(A-T)] as template shows that the enzyme copies the latter accurately. 4. The enzyme initiates new chains with purine nucleoside triphosphates exclusively. 5. The product obtained in the standard assay conditions contains some high mol.wt. (>16S) material, but consists primarily of short chains, of average length 70-80 nucleotide units. 6. The template specificity of the complete enzyme has been studied at high and low ionic strength. Its extreme dependence on salt concentration is unrelated to the gross overall base composition of the DNA used. 7. T(7) DNA is transcribed asymmetrically and the enzyme selectively copies the T(7) ;early' genes. 8. Preliminary amino acid analyses of alpha and beta subunits show that their overall content of acidic, basic and neutral amino acids does not differ appreciably from that of Escherichia coli RNA polymerase.
D000595:Amino Acid Sequence; D002247:Carbon Isotopes; D004247:DNA; D004586:Electrophoresis; D005815:Genetic Code; D006217:Halobacterium / Q000201:enzymology*; D008970:Molecular Weight; D009693:Nucleic Acid Hybridization; D009711:Nucleotides; D010759:Phosphorus Isotopes; D012316:RNA Nucleotidyltransferases / Q000302:isolation & purification*; D014316:Tritium
D002247:Carbon Isotopes; D009711:Nucleotides; D010759:Phosphorus Isotopes; D014316:Tritium; D004247:DNA; D012316:RNA Nucleotidyltransferases