Styrene-catabolism regulation in Pseudomonas fluorescens ST: phosphorylation of StyR induces dimerization and cooperative DNA-binding

Styrene is an important chemical extensively used in the petrochemical and polymer industries. In Pseudomonas fluorescens ST, styrene metabolism is controlled by a two-component regulatory system, very uncommon in the degradation of aromatic compounds. The two-component regulatory proteins StyS and StyR regulate the expression of the styABCD operon, which codes for styrene degradation. StyS corresponds to the sensor kinase and StyR to the response regulator, which is essential for the activation of PstyA, the promoter of the catabolic operon. In two-component systems, the response regulator is phosphorylated by the cognate sensor kinase. Phosphorylation activates the response regulator, inducing DNA-binding. The mechanism underlying this activation has been reported only for a very few response regulators. Here, the effect of phosphorylation on the oligomeric state and on the DNA-binding properties of StyR has been investigated. Phosphorylation induces dimerization of StyR, the affinity of dimeric StyR for the target DNA is higher than that of the monomer, moreover dimeric StyR binding to the DNA target is cooperative. Furthermore, StyR oligomerization may be driven by the DNA target. This is the first direct demonstration that StyR response regulator binds to the PstyA promoter.     

The styrene-responsive StyS/StyR regulation system controls expression of an auxiliary phenylacetyl-coenzyme A ligase: implications for rapid metabolic coupling of the styrene upper- and lower-degradative pathways

Pseudomonas sp. strain Y2 degrades styrene through oxidation to phenylacetic acid via the styABCD operon-encoded enzymes, whose expression is induced in response to styrene by the StyS/StyR two-component regulatory system. Further transformation of phenylacetic acid to tricarboxylic acid cycle intermediates is mediated by the enzymes of paa catabolic genes, whose expression is regulated by the PaaX repressor. The first step of this paa degradation pathway is catalysed by paaF-encoded phenylacetyl-coenzyme A ligases that produce phenylacetyl-coenzyme A. This metabolic intermediate, upon being bound by PaaX, inactivates PaaX-mediated repression of both the paa genes and the styABCD operon. Strain Y2 is unique in having three paaF genes located within two complete copies of the paa gene clusters. Expression of both paaF and paaF3 is controlled by the PaaX repressor. Here we use specific mutants in combination with in vivo and in vitro assays to demonstrate that paaF2, adjacent to the StyS/StyR regulatory genes, belongs to the StyR regulon and is not subject to repression by PaaX. We propose that this unexpected styrene-responsive regulatory strategy for the otherwise metabolically redundant PaaF2 auxiliary enzyme provides a system for rapid co-ordinate de-repression of the two sets of catabolic genes required for styrene degradation.     

Integration host factor amplifies the induction of the aceBAK operon of Escherichia coli by relieving IclR repression.

A binding site for integration host factor (IHF) was identified upstream of the aceBAK promoter. Under inducing conditions, IHF activates aceB::lacZ expression by opposing IclR repression. In contrast, IHF has little effect on aceB::lacZ expression under repressing conditions. The ability of IHF to relieve repression under inducing but not repressing conditions allows this protein to amplify the induction of aceBAK.