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Interaction of the Escherichia coli transporter DctA with the sensor kinase DcuS: presence of functional DctA/DcuS sensor units
Author(s) -
Witan Julian,
Bauer Julia,
Wittig Ilka,
Steinmetz Philipp Aloysius,
Erker Wolfgang,
Unden Gottfried
Publication year - 2012
Publication title -
molecular microbiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.857
H-Index - 247
eISSN - 1365-2958
pISSN - 0950-382X
DOI - 10.1111/j.1365-2958.2012.08143.x
Subject(s) - biology , biochemistry , escherichia coli , transmembrane domain , antiporter , transporter , microbiology and biotechnology , receptor , gene , membrane
Summary The aerobic Escherichia coli C 4 ‐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 Glt Ph , 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 Glt Ph . DctA contains an additional predicted amphipathic helix 8b on the cytoplasmic side of the membrane that is specific for DctA and not present in Glt Ph . 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 PAS C 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.