Conversion of the sensor kinase DcuS of Escherichia coli of the DcuB/DcuS sensor complex to the C₄‐dicarboxylate responsive form by the transporter DcuB

Abstract

The sensor kinase DcuS of Escherichia coli co-operates under aerobic conditions with the C -dicarboxylate transporter DctA to form the DctA/DcuS sensor complex. Under anaerobic conditions C -dicarboxylate transport in fumarate respiration is catalyzed by C -dicarboxylate/fumarate antiporter DcuB. (i) DcuB interacted with DcuS as demonstrated by a bacterial two-hybrid system (BACTH) and by co-chromatography of the solubilized membrane-proteins (mHPINE assay). (ii) In the DcuB/DcuS complex only DcuS served as the sensor since mutations in the substrate site of DcuS changed substrate specificity of sensing, and substrates maleate or 3-nitropropionate induced DcuS response without affecting the fumarate site of DcuB. (iii) The half-maximal concentration for induction of DcuS by fumarate (1 to 2 mM) and the corresponding K for transport (50 µM) differ by a factor of 20 to 40. Therefore, the fumarate sites are different in transport and sensing. (iv) Increasing levels of DcuB converted DcuS from the permanent ON (DcuB deficient) state to the fumarate responsive form. Overall, the data show that DcuS and DcuB form a DcuB/DcuS complex representing the C -dicarboxylate responsive form, and that the sensory site of the complex is located in DcuS whereas DcuB is required for converting DcuS to the sensory competent state.

Cellular Concentrations of the Transporters DctA and DcuB and the Sensor DcuS of Escherichia coli and the Contributions of Free and Complexed DcuS to Transcriptional Regulation by DcuR

“…With stronger induction (10 M arabinose), a significant increase (2.5-fold) was observed. These results are in line with data reported earlier (13).…”

“…DcuS requires one of the C 4 -dicarboxylate transporters DctA or DcuB for normal function. The transporters form a complex with DcuS, and only the DctA-DcuS and DcuB-DcuS complexes gain responsiveness to the C 4 -dicarboxylates, whereas free DcuS (DcuS F ) is in the permanent "on" state, where signaling occurs independent of C 4 -dicarboxylates (7,(11)(12)(13)15). A model was put forward that suggests two modes of regulation by DcuS (14).…”

“…These data were related to the function of DcuS in gene expression and of DcuB in transport. In particular, the responsiveness of DcuS to activation by fumarate serves as a measure for differentiating DcuS in the free or complexed state (12,13). The data shall show whether sufficient transporter is present in the noninduced state (absence of fumarate) for the formation of the sensory DcuB-DcuS (and DctA-DcuS) complexes, or whether there are indications for free (noncomplexed) DcuS (DcuS F ).…”

“…DcuS requires the transporter DctA or DcuB under aerobic and anaerobic conditions for sensing. The transporters form a complex with DcuS, converting DcuS to the C 4dicarboxylate-responsive state (11)(12)(13)(14). In this state, DcuS responds to and requires the C 4 -dicarboxylates for activation.…”

Transporters as information processors in bacterial signalling pathways

“…DctA therefore clearly acts as an activity switch for DcuS and does not contribute any additional sensory functions to the signalling process. It was recently shown that a functionally equivalent complex is formed between DcuS and DcuB under anaerobic conditions (Wörner et al ., ).…”