Nucleic Acids Research, 1990, Vol. 18, No. 10 2875-2880
© 1990
MOLECULAR BIOLOGY |
Quantitative analysis of Tn10 Tet repressor binding to complete set of tet operator mutants
Lehrstuhl für mikrobiologie. Friedrich Alexander Universität Erlangen/Nümberg Staudtrasse 5, D-8520 Erlangen, FRG
*To whom correspondence should be addressed
Received March 2, 1990. Accepted April 20, 1990.
A saturating ollgonucleotide-dlrected mutagenesis of both tet operators in the tet regulatory sequence was performed yielding mutants with four identical base pair exchanges at equivalent positions in the four tet operator half sides. The mutants were cloned between bipolar lacZ and galK indicator genes on a multicopy plasmid allowing the quantitative analysis of their effects in vivo. In the absence of Tet repressor the mutations lead to considerably different expression levels of both genes. They are discussed with respect to the promoter consensus sequences. In particular, the - 1 0 region of the in vivo active tetPR2 promoter is unambiguously defined by these results. In the presence of Tet repressor most of the mutants exhibit a lower affinity for that protein as determined quantitatively by their reduced expression levels. In general, tet operator recognition is most strongly affected by alterations of base pairs near the center of the palindromic sequence. The most important position is the third base pair, followed by base pairs two, four, five and six, the latter showing similar effects as base pair one. At each position, the four possible base pairs show different affinities for Tet repressor. They are discussed according to their exposure of H-bond donors and -acceptors in the major and minor grooves of the B-DNA. The results are in agreement with major groove contacts at positions two, three and five. At position four a low potential correlation of efficiencies with the H-bonding in the minor groove is found, while mutations at position six seem to influence repressor binding by other mechanisms.
CiteULike 
Connotea 
Del.icio.us What's this?
This article has been cited by other articles:
|
|
 |
|
  J. L. Ramos, M. Martinez-Bueno, A. J. Molina-Henares, W. Teran, K. Watanabe, X. Zhang, M. T. Gallegos, R. Brennan, and R. Tobes The TetR Family of Transcriptional Repressors Microbiol. Mol. Biol. Rev., June 1, 2005; 69(2): 326 - 356. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 A. Kamionka, M. Sehnal, O. Scholz, and W. Hillen Independent Regulation of Two Genes in Escherichia coli by Tetracyclines and Tet Repressor Variants J. Bacteriol., July 1, 2004; 186(13): 4399 - 4401. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 Y. Katan-Khaykovich and K. Struhl Dynamics of global histone acetylation and deacetylation in vivo: rapid restoration of normal histone acetylation status upon removal of activators and repressors Genes & Dev., March 15, 2002; 16(6): 743 - 752. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. Biburger, C. Berens, T. Lederer, T. Krec, and W. Hillen Intragenic Suppressors of Induction-Deficient TetR Mutants: Localization and Potential Mechanism of Action J. Bacteriol., February 1, 1998; 180(3): 737 - 741. [Abstract] [Full Text]
|
|
|
|
 |
|
 W Hinrichs, C Kisker, M Duvel, A Muller, K Tovar, W Hillen, and W Saenger Structure of the Tet repressor-tetracycline complex and regulation of antibiotic resistance Science, April 15, 1994; 264(5157): 418 - 420. [Abstract] [PDF]
|
|
|
|
 |
|
 W. Jiang, L. Zhou, B. Breyer, T. Feng, H. Cheng, R. Haydon, A. Ishikawa, and T.-C. He Tetracycline-regulated Gene Expression Mediated by a Novel Chimeric Repressor That Recruits Histone Deacetylases in Mammalian Cells J. Biol. Chem., November 21, 2001; 276(48): 45168 - 45174. [Abstract] [Full Text] [PDF]
|
|