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Mechanistic basis for RNA aptamer-based induction of TetR.

Steber, Markus ; Arora, Amit ; Hofmann, Jan ; Brutschy, Bernhard ; Suess, Beatrix (2011):
Mechanistic basis for RNA aptamer-based induction of TetR.
In: Chembiochem : a European journal of chemical biology, 12 (17), pp. 2608-14. ISSN 1439-7633,
[Article]

Abstract

The TetR aptamer induces TetR controlled gene expression, and represents an interesting tool for application in synthetic biology. We have analysed the mechanistic basis for RNA aptamer-based induction of TetR. The aptamer binds TetR with a high affinity in the order of 10(7)  M(-1), which is similar to operator DNA binding under the used ionic conditions. We identified the binding epitope of the aptamer on TetR, which consists of amino acids T27, N47 and K48 of both monomers, using loss-of-function analysis and electrophoretic mobility shift assays. Tetracycline-induced conformational changes of TetR led to reorientation of the DNA reading head. This movement destroys the composite binding epitope for the aptamer and leads to reduced RNA binding by one order of magnitude. The aptamer can actively displace TetR from the operator DNA; this could be the key factor for its activity in vivo.

Item Type: Article
Erschienen: 2011
Creators: Steber, Markus ; Arora, Amit ; Hofmann, Jan ; Brutschy, Bernhard ; Suess, Beatrix
Title: Mechanistic basis for RNA aptamer-based induction of TetR.
Language: English
Abstract:

The TetR aptamer induces TetR controlled gene expression, and represents an interesting tool for application in synthetic biology. We have analysed the mechanistic basis for RNA aptamer-based induction of TetR. The aptamer binds TetR with a high affinity in the order of 10(7)  M(-1), which is similar to operator DNA binding under the used ionic conditions. We identified the binding epitope of the aptamer on TetR, which consists of amino acids T27, N47 and K48 of both monomers, using loss-of-function analysis and electrophoretic mobility shift assays. Tetracycline-induced conformational changes of TetR led to reorientation of the DNA reading head. This movement destroys the composite binding epitope for the aptamer and leads to reduced RNA binding by one order of magnitude. The aptamer can actively displace TetR from the operator DNA; this could be the key factor for its activity in vivo.

Journal or Publication Title: Chembiochem : a European journal of chemical biology
Journal volume: 12
Number: 17
Divisions: 10 Department of Biology > Synthetic Genetic Circuits
10 Department of Biology
Date Deposited: 22 Feb 2012 10:11
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