Overproduction and purification of sigma 32, the Escherichia coli heat shock transcription factor

Protein Expr Purif. 1993 Oct;4(5):425-33. doi: 10.1006/prep.1993.1056.

Abstract

This paper reports the overproduction and the details of a rapid method to purify active sigma 32 that is free of core RNA polymerase enzyme. Maximal overproduction of sigma 32 in a T7 RNA polymerase-based expression system is achieved only in the presence of rifampicin. This 2-day procedure involves solubilizing inclusion bodies in Sarkosyl, removal of Sarkosyl by dialysis, and a single S-Sepharose column chromatography step. The final yield of sigma 32 is about 4.1 mg of approximately 95% purity from 1 g of wet weight cells.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Chromatography, Agarose
  • Escherichia coli / genetics*
  • Gene Expression
  • Genetic Vectors
  • Heat-Shock Proteins / biosynthesis*
  • Heat-Shock Proteins / genetics
  • Heat-Shock Proteins / isolation & purification
  • Recombinant Fusion Proteins / biosynthesis*
  • Recombinant Fusion Proteins / genetics
  • Recombinant Fusion Proteins / isolation & purification
  • Rifampin / pharmacology
  • Sigma Factor / biosynthesis*
  • Sigma Factor / genetics
  • Sigma Factor / isolation & purification
  • Transcription Factors*
  • Transcription, Genetic

Substances

  • Heat-Shock Proteins
  • Recombinant Fusion Proteins
  • Sigma Factor
  • Transcription Factors
  • heat-shock sigma factor 32
  • Rifampin