The role of Halobacterium cutirubrum deoxyribonucleic acid-dependent ribonucleic acid polymerase subunits in initiation and polymerization

1. The two subunits α and β of Halobacterium cutirubrum DNA-dependent RNA polymerase are required in equimolar amounts for RNA synthesis to occur in vitro at the maximum rate. 2. In the absence of bivalent cations no interaction occurs between α and β subunits or between the subunits and DNA. 3. Mn²⁺ causes the subunits to form a 1:1 complex that still does not bind to the template. 4. Mg²⁺ permits binding of the Mn²⁺-mediated complex to DNA. 5. The complete enzyme, αβ, is inhibited by rifampicin and only the β subunit relieves the inhibition when added in excess. 6. Rifampicin-insensitive, template-dependent RNA synthesis occurs in the presence of protein α alone provided an oligonucleotide with a 5′-purine terminus is supplied as primer. 7. In the primed reaction with the α protein and an oligonucleotide, the template specificity is independent of the ionic strength, in contrast with the marked effect of salt concentration on the template specificity of the complete enzyme. 8. It is concluded that the β protein controls the specificity of chain initiation and the template specificity of the complete enzyme and also carries the rifampicin-binding site, whereas the catalytic site is on the α subunit.