Structures of the ON and OFF states of the membraneous fumarate/Citrate sensor kinases DcuS from E. coli and CitA from Geobacillus thermodenitrifcans
Final Report Abstract
Transmembrane signaling by membrane-bound sensor kinases of bacteria which represent a large and diverse group of sensors is still enigmatic. Atomic resolution was obtained earlier for the nitrate sensor NarQ for a fragment consisting of the four-helical nitrate binding domain, the two TM helices and a cytoplasmic HAMP domain, but lacking the kinase core. This work gave ideas on the function of a specific class of TM sensor kinases. The mechanism of TM signaling by the CitA/DcuS family of sensor kinases was characterized in high resolution by the groups of this consortium in the present and earlier work. This class of sensor kinases is triggered by compaction of effector binding to the periplasmic PASP domain, followed by piston TM signaling, and by signal conversion on the cytoplasmic side by the cytosolic Linker/PASC domains. Details of signaling by PASC and signal transfer to the kinase have to be solved. The PASC dimer switching described here for Gt-CitAhas not been experimentally observed for any member of sensor kinases previously and could be the mechanism for kinase activation for a large number of representatives of this protein class. The work relied on the collaboration of groups using two complementary members of the CitA/DcuS family which allowed complementary studies on full length protein in situ (DcuS) and purified preparations of various constructs of the sensor (mostly CitA) by large set of experimental approaches. While the reported NMR work on CitA was done in the Griesinger department with the solid state NMR expertise of Dr. Loren Andreas, functional assays of the CitA mutants were performed in the Unden group. The PELDOR (collaboration with Hinderberger group) produced results for the switching in PASC of DcuS but the MTSL labels disturbed the enzymatic activity for Gt-CitA. Fortunately, CODEX (not described in the original project application) labels allowed the detection of the switching of the PASC domains of Gt-CitA. Biochemical and molecular genetic studies with DcuS enabled studies on the full-length DcuS protein in situ and in a fully functional context, yet with lower resolution. Discrepancies between the models of the signal transfer in the PASC region of DcuS and CitA are likely based on functional differences (DcuS requires the transporter DctA as a structural component of the sensor complex), or on the type of constructs.
Publications
- (2021) Conversion of the sensor kinase DcuS to the fumarate sensitive state by interaction of the bifunctional transporter DctA at the TM2/PASC-linker region. Microorganisms. 9(7), 1397
Stopp M, Schubert C, Unden G
(See online at https://doi.org/10.3390/microorganisms9071397) - (2021) Properties of transmembrane helix TM1 of the DcuS sensor kinase of Escherichia coli as the stator for TM2 piston signaling. Biol Chem, 402:1239-1246
Stopp M, Steinmetz PA, Unden G
(See online at https://doi.org/10.1515/hsz-2021-0254) - (2021) Transmembrane signaling and cytoplasmic signal conversion by dimeric transmembrane helix 2 and a linker domain of the DcuS sensor kinase. J Biol Chem.
Stopp M, Steinmetz PA, Schubert C, Griesinger C, Schneider D, Unden G
(See online at https://doi.org/10.1074/jbc.ra120.015999)
