Chapter 19 : Bacterial Toxin-Antitoxin Systems as Targets for the Development of Novel Antibiotics

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This chapter focuses on the proteic toxin-antitoxin (TA) systems. TA systems function as vitally important regulatory systems in bacteria and represent ideal targets for the development of novel antibiotic therapeutic agents. A broad mechanistic understanding of TA systems at physiological, biochemical, biophysical, and structural levels provides the scientific framework needed both for rational drug design and for elegant selection schemes using large pools of compounds. The proteins of chromosome-encoded TA systems (, , and ) from gram-negative bacteria, namely, CcdA-CcdB, Phd-Doc, ParD-ParE, YefM-YoeB, and one system from a plasmid from a G+ bacterium, have been studied in vitro with respect to their properties in solution and binding to DNA. The chapter summarizes the knowledge accumulated on these proteins. Pathogenic bacteria are subjected to an enormous selective pressure because of the indiscriminate overuse and misuse of broad-spectrum antibiotics. The recognition of the importance of protein-protein interactions within the cell has led to their investigation as targets for novel inhibitors. Here, the approaches that can be used for screening of inhibitors of protein-protein interactions are highlighted by recent research on the TA systems. The chapter focuses on two resonance energy transfer techniques, namely, fluorescent resonance energy transfer (FRET) and, especially, bioluminescence resonance energy transfer (BRET), since they have demonstrated to be highly useful for studying interactions between two proteins that have been shown to form complexes.

Citation: Alonso J, Balsa D, Cherny I, Christensen S, Espinosa M, Francuski D, Gazit E, Gerdes K, Hitchin E, Martín M, Nieto C, Overweg K, Pellicer T, Saenger W, Welfle H, Welfle K, Wells J. 2007. Bacterial Toxin-Antitoxin Systems as Targets for the Development of Novel Antibiotics, p 313-329. In Bonomo R, Tolmasky M (ed), Enzyme-Mediated Resistance to Antibiotics. ASM Press, Washington, DC. doi: 10.1128/9781555815615.ch19
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Image of Figure 19.1
Figure 19.1

(Right) Three-dimensional structures of homodimeric toxins Kid and CcdB; α-helices are shown as spirals and β-strands are shown as arrows (7, 20; PDB codes 1VUB [ ] and 1M1F [ ]). The noncrystallographic twofold axes relating the monomers are vertical to the paper plane and indicated by ellipses, β-strands are numbered, N and C termini are labeled, and a and b mark the positions where loops are disordered and not seen in the electron density. (Left) The TA complex MazF-MazE-MazF (PDB code 1UB4 [ ]) has twofold crystallographic symmetry indicated by the ellipse in the center of MazE. The orientation of the two MazF is similar to that of CcdB and Kid in order to illustrate their structural homology. β-Strands of MazF and MazE are numbered, and α-helices of MazE are labeled α1 and α2; α-helices and loops of MazE are drawn darker than for MazF. The MazE C termini (one is labeled C) bind to the two MazF homodimers. The loop between β-strands β1 and β2 in MazF is disordered (labeled d) and not seen in the electron density. The sequence of α-helices and β-strands is β1-β2-α1-β3-β5-β6-α2-β7-α3 in Kid and MazF, and in CcdB α1 is shifted and located between β5 and β6. For MazE the sequence is β1-β2-α1-β3-β4-α2.

Citation: Alonso J, Balsa D, Cherny I, Christensen S, Espinosa M, Francuski D, Gazit E, Gerdes K, Hitchin E, Martín M, Nieto C, Overweg K, Pellicer T, Saenger W, Welfle H, Welfle K, Wells J. 2007. Bacterial Toxin-Antitoxin Systems as Targets for the Development of Novel Antibiotics, p 313-329. In Bonomo R, Tolmasky M (ed), Enzyme-Mediated Resistance to Antibiotics. ASM Press, Washington, DC. doi: 10.1128/9781555815615.ch19
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Image of Figure 19.2
Figure 19.2

(Top) Topography of ε and ζ. α-Helices are indicated by circles and labeled a to c in ε and A to L in ζ. The polarity of β-strands (large numbers) is given by up and down pointing triangles, and small numbers mark positions in amino acid sequences. (Bottom) Three-dimensional dumbbell-shaped structure of complex εζ (PDB code 1gvn). α-Helices and β-strands are labeled as in the top panel. The noncrystallographic twofold axis relating εζ dimers in the heterotetrameric εζ complex is indicated by a vertical line; termini are labeled N, C, and C′.

Citation: Alonso J, Balsa D, Cherny I, Christensen S, Espinosa M, Francuski D, Gazit E, Gerdes K, Hitchin E, Martín M, Nieto C, Overweg K, Pellicer T, Saenger W, Welfle H, Welfle K, Wells J. 2007. Bacterial Toxin-Antitoxin Systems as Targets for the Development of Novel Antibiotics, p 313-329. In Bonomo R, Tolmasky M (ed), Enzyme-Mediated Resistance to Antibiotics. ASM Press, Washington, DC. doi: 10.1128/9781555815615.ch19
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Image of Figure 19.3
Figure 19.3

Putative active site of ζ ( ). Functional amino acids are drawn and labeled, ATP and the essential Mg are modeled according to the structure of Cmp, and the binding site for the yet unknown substrate is indicated by an ellipse.

Citation: Alonso J, Balsa D, Cherny I, Christensen S, Espinosa M, Francuski D, Gazit E, Gerdes K, Hitchin E, Martín M, Nieto C, Overweg K, Pellicer T, Saenger W, Welfle H, Welfle K, Wells J. 2007. Bacterial Toxin-Antitoxin Systems as Targets for the Development of Novel Antibiotics, p 313-329. In Bonomo R, Tolmasky M (ed), Enzyme-Mediated Resistance to Antibiotics. ASM Press, Washington, DC. doi: 10.1128/9781555815615.ch19
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Table 19.1

TA systems identified on plasmids and chromosomes

Citation: Alonso J, Balsa D, Cherny I, Christensen S, Espinosa M, Francuski D, Gazit E, Gerdes K, Hitchin E, Martín M, Nieto C, Overweg K, Pellicer T, Saenger W, Welfle H, Welfle K, Wells J. 2007. Bacterial Toxin-Antitoxin Systems as Targets for the Development of Novel Antibiotics, p 313-329. In Bonomo R, Tolmasky M (ed), Enzyme-Mediated Resistance to Antibiotics. ASM Press, Washington, DC. doi: 10.1128/9781555815615.ch19

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