Purification and properties of ribonuclease E, an RNA-processing enzyme from Escherichia coli

Abstract

The Escherichia coli RNA-processing enzyme RNAase E was purified through a number of steps including isoelectrofocusing. The final fraction contained mainly a single polypeptide of 66 kDa. However, while all the steps in the purification yielded the same qualitative activity, the specific activity of fractions was decreased in the last steps of the purification. By combining the most-purified enzyme with earlier fractions from the purification, we could show that the cells could contain a factor that enhances RNAase E activity. The purified enzyme showed the same characteristics with respect to temperature optimum, pH and ionic requirements as less-purified preparations. Testing specific inhibitors we concluded that the enzyme requires SH groups, free amino groups, and either of the amino acids tryptophan, tyrosine, histidine or methionine for its activity.