Abstract
Bacillus subtilis core RNA polymerase, containing a His(6)-fusion to the C-terminus of the beta' subunit, was isolated by Ni-NTA, Superdex 200 gel filtration, and Mono Q anion-exchange chromatography. The purified core enzyme was shown to be free of the major sigma factor(A) and the transcription factors NusA and GreA. The purification procedure can be completed within 1 working day, is scalable, and yields highly purified and active core RNA polymerase.
Copyright 2000 Academic Press.
Publication types
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Research Support, U.S. Gov't, Non-P.H.S.
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Research Support, U.S. Gov't, P.H.S.
MeSH terms
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Bacillus subtilis / genetics
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Bacillus subtilis / metabolism*
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Chromatography
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Chromatography, Affinity
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DNA-Directed RNA Polymerases / genetics*
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DNA-Directed RNA Polymerases / isolation & purification*
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DNA-Directed RNA Polymerases / metabolism
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Electrophoresis, Polyacrylamide Gel
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Enzyme Activation
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Gene Expression Regulation, Bacterial / physiology
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Molecular Probes
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Recombinant Fusion Proteins / genetics*
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Recombinant Fusion Proteins / isolation & purification*
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Recombinant Fusion Proteins / metabolism
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Transcription Factors / metabolism
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Transcription, Genetic
Substances
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Molecular Probes
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Recombinant Fusion Proteins
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Transcription Factors
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beta' subunit of RNA polymerase
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DNA-Directed RNA Polymerases