Procedure for the rapid, large-scale purification of Escherichia coli DNA-dependent RNA polymerase involving polymin P precipitation and DNA-cellulose chromatography

Abstract

An improved method is described for the purification of the DNA-dependent RNA polymerase [ribonucleosidetriphosphate:RNA nucleotidyltransferase, EC 2.7.7.6] from Escherichia coli. The method involves lysozyme-sodium deoxycholate lysis, low-speed centrifugation, precipitation with Polymin P, elution from the Polymin P precipitate, ammonium sulfate precipitation, and chromatography on DNA-cellulose and Bio-Gel A 5m. RNA polymerase is purified to electrophoretic homogeneity in 2 days with a recovery of 45%, resulting in a yield of 250 mg of holoenzyme from 500 g of cells.