PMID: 2111806Jun 1, 1990Paper

Overproduction, purification, and characterization of Bacillus subtilis RNA polymerase sigma A factor

Journal of Bacteriology
B Y Chang, R H Doi

Abstract

By use of a T7 expression system, large amounts of active Bacillus subtilis RNA polymerase sigma A factor were produced in Escherichia coli cells. This overproduced protein was found in the form of inclusion bodies and constituted 40% of the total cellular protein. Because of the ease of isolation of the inclusion bodies and the acidic properties of sigma A, the protein was purified to more than 99% purity and the yield was about 90 mg/liter of culture. Gel mobility, antigenicity, specificity of promoter recognition, and N-terminal amino acid sequence of the overproduced sigma were found to be the same as those of native sigma A. Partial proteolysis analysis of sigma A protein suggested the presence of a protease-sensitive surface region in the C-terminal part of the sigma A protein. The promoter -10 binding region of sigma A was less sensitive to proteases and was probably involved in a hydrophobic, tightly folded domain of sigma A protein.

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Citations

Oct 12, 2010·Nucleic Acids Research·Hsin-Yi YehBan-Yang Chang
Mar 24, 2016·Nucleic Acids Research·Veronika RaindlováMichal Hocek
Nov 13, 2001·Molecular Microbiology·J A OpdykeC P Moran
Sep 6, 1996·The Journal of Biological Chemistry·R Deora, T K Misra
May 21, 1999·The Journal of Biological Chemistry·F J López de SaroJ D Helmann
Aug 5, 1998·Proceedings of the National Academy of Sciences of the United States of America·J H KimG H Chambliss
Dec 17, 2019·The EMBO Journal·Michaela ŠikováLibor Krásný
Feb 13, 2014·Nucleic Acids Research·Jana WiedermannováLibor Krásny
Jan 7, 2021·Microorganisms·Petra SudzinováLibor Krásný
Mar 1, 1995·Microbiological Reviews·W G Haldenwang
Jun 1, 1992·Journal of Bacteriology·M LonettoC A Gross
Feb 26, 2008·Journal of Bacteriology·Amrita Kumar, Charles P Moran
Aug 20, 2004·Journal of Bacteriology·Kuei-Min ChungBan-Yang Chang
Feb 22, 2008·Protein Expression and Purification·Xiao Yang, Peter J Lewis
Oct 25, 2008·Methods : a Companion to Methods in Enzymology·Irina Artsimovitch, Tina M Henkin
Sep 11, 2021·Journal of Microbiological Methods·Sweetha GanapathyRoland K Hartmann

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