Interaction of the Escherichia coli transporter DctA with the sensor kinase DcuS: presence of functional DctA/DcuS sensor units

Abstract

Summary The aerobic Escherichia coli C4‐dicarboxylate transporter DctA and the anaerobic fumarate/succinate antiporter DcuB function as obligate co‐sensors of the fumarate responsive sensor kinase DcuS under aerobic or anaerobic conditions respectively. Overproduction under anaerobic conditions allowed DctA to replace DcuB in co‐sensing, indicating their functional equivalence in this capacity. In vivo interaction studies between DctA and DcuS using FRET or a bacterial two‐hybrid system (BACTH) demonstrated their interaction. DctA–YFP bound to an affinity column and was able to retain DcuS. DctA shows substantial sequence and secondary structure conservation to GltPh, the Na+/glutamate symporter of Pyrococcus horikoshii with known 3D structure. Topology studies of DctA demonstrated the presence of eight transmembrane helices in an arrangement similar to that of GltPh. DctA contains an additional predicted amphipathic helix 8b on the cytoplasmic side of the membrane that is specific for DctA and not present in GltPh. Mutational analysis demonstrated the importance of helix 8b in co‐sensing and interaction with DcuS, and the isolated helix 8b showed strong interaction with DcuS. In DcuS, deletion and mutation of the cytoplasmic PASC domain affected the interaction between DctA and DcuS. It is concluded that DctA forms a functional unit or sensor complex with DcuS through specific interaction sites.

The Sensor Kinase DctS Forms a Tripartite Sensor Unit with DctB and DctA for Sensing C4-Dicarboxylates in Bacillus subtilis

“…7), confirming the functional interdependence of DctA and DctS in B. subtilis (13). The physical and functional interaction suggests that DctA functions as a cosensor of DctS similarly to the DctA/DcuS complex in E. coli (12,33). DctA of E. coli interacts with DcuS by the C-terminal ␣-helix 8b (12).…”

“…In addition, DcuS requires the C 4 -dicarboxylate transporter DctA or DcuB for function, and possibly also DauA plays an essential role in sensing by DcuS; in dctAor dcuB-deficient strains, DcuS is deregulated and in the permanent ON state even in the absence of C 4 -dicarboxylates (10,11). DcuS and the transporter DctA interact physically, suggesting the formation of a DctA/DcuS sensor complex (12).…”

Identification of a Novel Nutrient-Sensing Histidine Kinase/Response Regulator Network in Escherichia coli

“…Interactions between sensory proteins and transporters, the so-called trigger transporter mechanism, have already been described (25). Well-characterized examples include the bacitracin resistance module BceS/BceAB in Bacillus subtilis (26) and the cosensory systems DctA/DcuS (27) and CadC/LysP in E. coli (15).…”

Role of the Transporter-Like Sensor Kinase CbrA in Histidine Uptake and Signal Transduction

“…For SKs involved in nutrient acquisition, it seems reasonable to assume that specific binding of the nutrient molecule might enable transmission of the nutrient signal into the cytoplasm, leading to activation of genes for subsequent uptake (and degradation, if applicable). Indeed, such an arrangement has been demonstrated in the control and transport of C 4 dicarboxylates (13)(14)(15). For example, in the plant symbiotic bacteria Sinorhizobium meliloti (16) and Rhizobium leguminosarum (17), uptake of succinate is mediated by the DctA permease, whose expression is regulated by the two-component system DctB and DctD.…”