Ribonucleotide Reductase from Escherichia coli: Demonstration of a Highly Active Form of the Enzyme

Abstract

Ribonucleotide reductase from Escherichia coli consists of two nonidentical subunits, proteins B1 and B2. The activity of the enzyme in crude extracts prepared from mechanically disrupted bacteria is very low. Enzyme activity is stimulated 5 to 10-fold by addition of an excess of either subunit. Concentrated extracts from cells lysed gently on Cellophane discs (Schaller et al.) contained 10 to 20-fold higher activity than extracts from mechanically disrupted cells. This activity was not further stimulated by either B1 or B2. The system is suitable for complementation tests for the analysis of temperature-sensitive mutants affecting the ribonucleotide reductase system. Concentrated high-spced supernatants from E. coli treated with lysozyme (Wickner et al.) also contained a high ribonucleotide reductase activity, which was stimulated slightly or not at all by addition of B1 and B2. This active form of the enzyme was unstable and could not be purified. The results suggest that the intracellular form of the enzyme consists of a tight complex of proteins B1 and B2, possibly stabilized by other intracellular structures.