Sensory domain contraction in histidine kinase CitA triggers transmembrane signaling in the membrane-bound sensor

Abstract

Bacteria use membrane-integral sensor histidine kinases (HK) to perceive stimuli and transduce signals from the environment to the cytosol. Information on how the signal is transmitted across the membrane by HKs is still scarce. Combining both liquid-and solid-state NMR, we demonstrate that structural rearrangements in the extracytoplasmic, citrate-sensing Per-Arnt-Sim (PAS) domain of HK CitA are identical for the isolated domain in solution and in a longer construct containing the membrane-embedded HK and lacking only the kinase core. We show that upon citrate binding, the PAS domain contracts, resulting in a shortening of the C-terminal β-strand. We demonstrate that this contraction of the PAS domain, which is well characterized for the isolated domain, is the signal transmitted to the transmembrane (TM) helices in a CitA construct in liposomes. Putting the extracytoplasmic PAS domain into context of the membrane-embedded CitA construct slows down citrate-binding kinetics by at least a factor of 60, confirming that TM helix motions are linked to the citrate-binding event. Our results are confirmation of a hallmark of the HK signal transduction mechanism with atomic resolution on a full-length construct lacking only the kinase core domain.transmembrane receptors | solid-state NMR spectroscopy | transmembrane signaling | integrated structural biology | histidine sensor kinase H istidine kinases (HKs) are key players in prokaryotic signaling and the primary means of processing extracellular stimuli (1-6). Due to their modular organization, HKs can relay a multitude of different input stimuli, including sensing of nutrients, light, physicochemical parameters (e.g., metal ions, ions gradient), temperature, oxygen, and molecules reporting cell density, to accomplish diverse responses. Over the last decade, various structures have been solved for the cytosolic kinase core and a number of signaling domains and more recently even for the complete cytoplasmic region (7-13), but the transmembrane (TM) signaling mechanism itself remains challenging to decipher. Structural information on individual stimulus-receiving domains therefore provides an important tool to identify TM signal transduction schemes.Because extracytoplasmic receiver domains are generally directly connected to TM helices, structural rearrangements within these motifs can be correlated with the TM-signaling mechanism. Different receiver domains have been characterized with and without their ligands and reveal structural differences between the two states that impose restraints on possible motions of the TM helices. Based on such structures, a symmetry-to-asymmetry switch has been postulated for KinB (14), TorT/TorS (15), and LuxPQ (16), implying a possible tilt of TM helices (17). For chemotaxis sensor Tar and for HK NarX a piston movement of the second TM helix has been postulated as a consequence of receiver domain motions (9,(18)(19)(20).Among receiver domains, PAS domains stand out by being able to process diverse input signals, which is reflected by...

Transmembrane Signal Transduction in Two‐Component Systems: Piston, Scissoring, or Helical Rotation?

“…The model has also found support in simulations . Besides Tar, the experiments have also shown piston‐like motions in HKs CitA and DcuS . The helical rotation model is based on the observation of highly asymmetric ligand‐bound conformation of LuxQ periplasmic sensor domain .…”

“…Piston‐like motion, accompanied by helical rotations, were also identified later in α‐helical sensors of histidine kinases NarQ, NarX, and TorS; the latter binds its ligand via an accessory protein (TorT) . Contractions, which could cause piston‐like motions, were observed in single Cache sensor of histidine kinase CitA or double Cache sensor of Campylobacter jejuni chemoreceptor Tlp3 …”

Protein resonance assignment by BSH‐CP‐based 3D solid‐state NMR experiments: A practical guide

“…BSH‐CP‐based experiments are suited for a wide range of samples and have already been successfully applied to different kinds of proteins such as membrane proteins, amyloid fibrils, and microcrystalline protein samples, as well as protein filaments and supramolecular assemblies …”

High performance intensiometric direct- and inverse-response genetically encoded biosensors for citrate

“…Linked to TM2 are several intracellular domains, including the C-terminal histidine kinase (HK) catalytic domain 21 . Necessarily, binding of a cognate ligand to the periplasmic sensory domain leads to a conformational change that propagates across the membrane and modulates the activity of the C-terminal HK domain [21][22][23] . Various models have been proposed for these membrane-spanning structural changes including the "piston" model, in which the two helices are vertically displaced relative to each other, the "scissor blade" model, in which the helices undergo a diagonal scissor-like displacement, and the "rotating helix" model in which one helix is proposed to rotate relative to the other 21 .…”

“…One relatively well characterized SHK is the citrate-responsive CitA protein from Klebsiella pneumoniae 22,24,25 . X-ray crystal structures are available for the Per-Arnt-Sim (PAS)-fold citrate-binding periplasmic domain (CitAP) both with (PDB IDs 1P0Z 24 and 2J80) 25 and without (PDB ID 2V9A) 25 citrate ( Fig.…”