Phage Q-beta ribonucleic acid replicase. Subunit relationships determined by intramolecular cross-linking

Abstract

Phage Qbeta replicase is composed of four subunits of molecular weights 70,000, 65,000, 45,000, and 35,000. Treatment of the enzyme with protein cross-linking reagents results in formation of three covalently bound complexes of molecular weights 215,000, 135,000, and 80,000. Analysis of the two larger complexes formed by cross-linking with a reversible cross-linker (methyl-4-mercaptobutyrimidate) demonstrates that the 215,000 molecular weight complex is composed of one each of the replicase subunits, while the 135,000 molecular weight complex is composed of the two larger subunits. The 80,000 molecular weight complex was shown to be made up of the two smaller subunits by cross-linking these two subunits in the absence of the larger pair. Increasing ionic strength stabilizes the large complex at the expense of the two smaller complexes. The presence of stoichiometric amounts of Qbeta RNA during cross-linking dramatically reduces formation of the large complex; other natural and synthetic RNAs reduce the formation of this complex to a lesser extent.