## 5 Phd-Doc, RelB-RelE and Kis-Kid: Transcriptional Regulation and Conditional Cooperativity  
The lysogenic form of bacteriophage P1 exists as a low copy number plasmid that is stabilized by the Phd-Doc type II TA. The Doc toxin possesses high and low affinity binding sites for the Phd antitoxin. The Phd-Doc complex is a highly effective transcriptional repressor when both sites are occupied, whereas Phd alone represses only weakly [63,64]. If the Doc:Phd ratio increases sufficiently, the antitoxin molecules are captured preferentially by the high affinity binding sites in the excess Doc proteins. The consequent alteration in quaternary structure of the Phd-Doc complex disrupts the repressive complex thereby inhibiting operator site binding. This leads to derepression of the _phd-doc_ operon specifically when toxin levels are elevated. More Phd is produced as a consequence to allow both trapping of excess Doc and re-establishment of transcriptional repression [64]. This phenomenon in which TA operon expression is modulated by the toxin:antitoxin ratio is termed _conditional cooperativity_[65,66] and reflects earlier observations that excess toxin often perturbed operator binding by TA complexes [37,44,67,68,69]. Conditional cooperativity is thought to calibrate TA levels so that random induction of operon expression is curtailed and also to permit efficient sequestration of excess toxin so that toxin-induced temporary stasis can be reversed [70]. RelB-RelE is one of the most prevalent and well-studied TA complexes [71, 72]. Like MazF, the RelE toxin is an endoribonuclease, albeit with a different cleavage specificity on mRNA [73, 74, 75]. Moreover, unlike MazF, transcript cleavage by RelE is translation-dependent [74]. The N-terminus of the RelB antitoxin adopts a dimeric RHH fold connected to a pair of C-terminal extensions that capture two RelE monomers, forming an open V-shaped heterotetramericic stucture in which RelB is positioned at the vertex [65, 76, 77]. This configuration blocks access by the toxin to the ribosomal A site and induces structural rearrangements that perturb the endoribonuclease pocket [65, 78]. Transcription of the _relB-relE_ module is autoregulated in _E. coli._ In common with numerous TA complexes, the RelB antitoxin represses transcription weakly whereas the RelE toxin is a co-repressor [79]. The operator site recognized by RelB-RelE comprises a pair of tandem 12-bp inverted repeats one of which overlaps the \(-10\) promoter motif for the _relBE_ module (Figure 3) [76, 77]. The RelB-RelE complex exhibits conditional cooperativity: RelE promotes operator binding by RelB at subequimolar ratios but ablates binding at higher concentrations [66]. In agreement, modelling of the complex onto DNA suggests that a RelB dimer associated with a single RelE monomer binds one inverted repeat within the operator. The second inverted repeat is occupied by another heterotrimeric complex that interacts cooperatively with the adjacent complex. In contrast, steric clashes between RelE molecules are predicted to block operator binding by a complex with an equimolar RelB:RelE ratio [65]. The outcome of conditional cooperativity in the RelB-RelE system parallels that of the Phd-Doc complex described above [64], _i.e._, differential regulation of the cognate TA genes by toxin:antitoxin ratios. As with Phd-Doc, RelE possesses high and low affinity binding sites for RelB that likely influence the RelB:RelE ratio intracellularly. The low affinity site for the RelB-RelE interaction could play a role in the interaction of excess of toxin with the TA repressor bound to the operator and in the release of the repressor under these circumstances. However, unlike with Phd-Doc, these sites are not invoked in conditional cooperativity. Instead, interactions between RelB dimers positioned on the inverted repeats and stabilized by subequimolar binding of RelE are sufficient for cooperative operator recognition [65]. The Kis-Kid type II TA complex is encoded by the R1 plasmid of _E.