This Article
Right arrow Full Text
Right arrow Full Text (PDF)
Right arrow Supplemental material
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Services
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrowReprints and Permissions
Right arrow Copyright Information
Right arrow Books from ASM Press
Right arrow MicrobeWorld
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Gößringer, M.
Right arrow Articles by Hartmann, R. K.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Gößringer, M.
Right arrow Articles by Hartmann, R. K.

 Previous Article  |  Next Article 

Journal of Bacteriology, October 2006, p. 6816-6823, Vol. 188, No. 19
0021-9193/06/$08.00+0     doi:10.1128/JB.00756-06
Copyright © 2006, American Society for Microbiology. All Rights Reserved.

Analysis of RNase P Protein (rnpA) Expression in Bacillus subtilis Utilizing Strains with Suppressible rnpA Expression{dagger}

Markus Gößringer,1 Rosel Kretschmer-Kazemi Far,2 and Roland K. Hartmann1*

Institut für Pharmazeutische Chemie, Philipps-Universität Marburg, Marbacher Weg 6, D-35037 Marburg, Germany,1 Universität zu Lübeck, Institut für Molekulare Medizin, Ratzeburger Allee 160, 23538 Lübeck, Germany2

Received 26 May 2006/ Accepted 21 July 2006

Bacterial RNase P is composed of an RNA subunit and a single protein subunit (encoded by the rnpB and rnpA genes, respectively). We constructed Bacillus subtilis mutant strains that conditionally express the RNase P protein under control of the xylose promoter (Pxyl). In one strain (d7), rnpA expression was efficiently repressed in the absence of the inducer xylose, leading to cell growth arrest. Growth could be restored by a second functional rnpA allele. This is the first RNase P protein knockdown strain, providing the first direct proof that the rnpA gene is essential in B. subtilis and, by inference, in other bacteria. We further show (i) that, in the wild-type context, rnpA expression is attenuated by transcriptional polarity and (ii) that translation of rnpA mRNA in B. subtilis can be initiated at two alternative start codons. His-tagged RNase P protein variants are functional in vivo and permit purification of in vivo-assembled holoenzymes by affinity chromatography. Simultaneous expression of plasmid-encoded RNase P RNA and His-tagged protein increased RNase P holoenzyme yields. Massive overproduction of RNase P protein in strain d7 is compatible with cell viability.

* Corresponding author. Mailing address: Institut für Pharmazeutische Chemie, Philipps-Universität Marburg, Marbacher Weg 6, D-35037 Marburg, Germany. Phone: 49-6421-2825827. Fax: 49-6421-2825854. E-mail: roland.hartmann{at}staff.uni-marburg.de.

{dagger} Supplemental material for this article may be found at http://jb.asm.org/.

Journal of Bacteriology, October 2006, p. 6816-6823, Vol. 188, No. 19
0021-9193/06/$08.00+0     doi:10.1128/JB.00756-06
Copyright © 2006, American Society for Microbiology. All Rights Reserved.



This article has been cited by other articles:

  • Hsieh, J., Walker, S. C., Fierke, C. A., Engelke, D. R. (2009). Pre-tRNA turnover catalyzed by the yeast nuclear RNase P holoenzyme is limited by product release. RNA 15: 224-234 [Abstract] [Full Text]  
  • Li, D., Willkomm, D. K., Hartmann, R. K. (2009). Minor changes largely restore catalytic activity of archaeal RNase P RNA from Methanothermobacter thermoautotrophicus. Nucleic Acids Res 37: 231-242 [Abstract] [Full Text]  
  • Marszalkowski, M., Willkomm, D. K., Hartmann, R. K. (2008). Structural basis of a ribozyme's thermostability: P1 L9 interdomain interaction in RNase P RNA. RNA 14: 127-133 [Abstract] [Full Text]  
  • Wegscheid, B., Hartmann, R. K. (2007). In vivo and in vitro investigation of bacterial type B RNase P interaction with tRNA 3'-CCA. Nucleic Acids Res 0: gkm005v2-14 [Abstract] [Full Text]  


Copyright © 2009 by the American Society for Microbiology.   For an alternate route to Journals.ASM.org, visit: http://intl-journals.asm.org | More Info»