DNA‐Dependent RNA Polymerase from Halobacterium halobium

Abstract

DNA‐dependent RNA polymerase core enzyme was isolated from Halobacterium halobium. The purification is based on the finding that the enzyme is stable in 40% (v/v) glycerol, in the presence of 0.05 M MgCl2 and involves adsorption of contaminants to DEAE‐cellulose, precipitation of the complex of polymerase with DNA by streptomycin sulfate, chromatography over Biogel and affinity chromatography over heparin‐Sepharose or heparin‐cellulose. The enzyme consists of four or five different subunits. The composition formula was estimated as (150 000) (86 000)2 (72 000)2 (49 000)3 or 2; there may be one or two different 49 000‐Mr subunits. RNA synthesis requires a template. Denatured DNA is more efficient than native DNA. The transcription of native DNA is specifically stimulated by the addition of a possibly σ‐like factor eluted from DEAE‐cellulose. The fidelity of transcription is indicated by the absolute requirement for UTP besides ATP with poly[d(A‐T)] as the template. Copyright © 1978, Wiley Blackwell. All rights reserved