The Tetracycline Repressor—A Paradigm for a Biological Switch

Abstract

The excessive use of antibiotics has enabled bacteria to develop resistance through a variety of mechanisms. The most common bacteriostatic action of the broad‐spectrum antibiotic tetracycline (Tc) is by the inactivation of the bacterial ribosome so that the protein biosynthesis is interrupted and the bacteria die. The most common mechanism of resistance in gram‐negative bacteria against Tc is associated with the membrane‐intrinsic protein TetA, which exports invaded Tc out of the bacterial cell before it can attack its target, the ribosome. The expression of TetA is tightly regulated by the homodimeric Tet repressor (TetR)2, which binds specifically with two helix‐turn‐helix motifs of operator DNA (tetO; Kass≈1011 M−1) located upstream from the tetA gene on a plasmid or transposon. When Tc diffuses into the cell it chelates Mg2+ and the complex [MgTc]+ binds to (TetR)2 to form the induced complex (TetR⋅[MgTc]+)2. This process is associated with conformational changes, which sharply reduce the affinity of (TetR)2 to tetO, so that expression of TetA can take place, thus conferring resistance to the bacteria cells against Tc. Crystallographic studies show sequence‐specific protein–nucleic acid interactions in the (TetR)2⋅tetO complex and how the binding of two [MgTc]+ to (TetR)2 enforces conformational changes that are stabilized by cooperative binding of two chains of eight water molecules each so that the formed (TetR⋅[MgTc]+)2 is no longer able to recognize and bind to tetO. Since the switching mechanisms of the TetR/[MgTc]+ system is so tight, it has proven very useful in the regulation of eukaryotic gene expression and may also be applicable in gene therapy.

Regulating transcription regulators via allostery and flexibility

“…Antibiotic binding to TetR leads to a decrease in affinity of the protein for DNA that allows transcription of the genes that code for TetA, a membrane protein that exports the tetracyline out of the bacterial cell before it can attack its target, the ribosome, and TetR itself (2). The repressor protein is a homodimer in which each monomer is composed of a tetracycline binding/dimerization domain and a helix-turn-helix DNA binding domain (3).…”

Synthesis, physico-chemical characterization and bacteriostatic study of Pt complexes with substituted amine ligands

“…These binding properties are studied and used in design and synthesis of new antibacterial drugs also [9,10]. As a high number of pathogenic bacteria show resistance toward antibiotics, it is necessary to find new effective compounds and control the multidrug resistance of microbes [11,12]. Therefore much effort is put into modifying of antibiotics already in use, while new compounds are also synthesized.…”

Drug-stabilized and drug-free complexes of topo IV fromS. pneumoniaeshed light on the mechanisms of reversible DNA scission and selective drug resistance

“…In all, at least 23 protein kinases are known to be phosphorylated by PDK1. We present the crystal structure [1] of PDK1 bound to a rationally developed low-molecular-weight activator [2] and describe the conformational changes induced by small compounds in the crystal and in solution using a fluorescencebased assay and deuterium exchange experiments. Our results indicate that the binding of the compound produces local changes at the target site, the PIF binding pocket, and also allosteric changes at the ATP binding site and the activation loop.…”

“…Altogether, we present molecular details of the allosteric changes induced by small compounds that trigger the activation of PDK1 through mimicry of phosphorylationdependent conformational changes. [1,2]. To receive further structural information on membrane thickness and the change of it by drug incorporation Small Angle X-Ray Scattering (SAXS) profiles have been recorded.…”

“…The "induced" complex (Tc-bound) is formed upon binding of Tc to TetR and causes the dissociation of TetR from tetO, so that the genes tetR (encoding TetR) and tetA (encoding antiporter TetA) responsible for Tc-resistance can be transcribed. The conformational switch between the two states has been characterized as a shift from a Tc-bound conformation with the DNA-binding domains locked in an orientation that is unfavourable for tetO binding to more flexible behaviour of the DNA-binding domains in the Tc-free state [1]. However, recently, several TetR mutants [2] and Tc peptide mimetics [3] have been identified that appear not to behave according to this "classical mechanism".…”