Allosteric mechanism of signal transduction in the two-component system histidine kinase PhoQ

Abstract

Transmembrane signaling proteins couple extracytosolic sensors to cytosolic effectors. Here, we examine how binding of Mg2+ to the sensor domain of an E. coli two component histidine kinase (HK), PhoQ, modulates its cytoplasmic kinase domain. We use cysteine-crosslinking and reporter-gene assays to simultaneously and independently probe the signaling state of PhoQ's sensor and autokinase domains in a set of over 30 mutants. Strikingly, conservative single-site mutations distant from the sensor or catalytic site strongly influence PhoQ's ligand-sensitivity as well as the magnitude and direction of the signal. Data from 35 mutants are explained by a semi-empirical three-domain model in which the sensor, intervening HAMP, and catalytic domains can adopt kinase-promoting or inhibiting conformations that are in allosteric communication. The catalytic and sensor domains intrinsically favor a constitutively 'kinase-on' conformation, while the HAMP domain favors the 'off' state; when coupled, they create a bistable system responsive to physiological concentrations of Mg2+. Mutations alter signaling by locally modulating domain intrinsic equilibrium constants and interdomain couplings. Our model suggests signals transmit via interdomain allostery rather than propagation of a single concerted conformational change, explaining the diversity of signaling structural transitions observed in individual HK domains.

Structural mechanism of signal transduction in a phytochrome histidine kinase

“…To the best of our knowledge, this is the first observation of such a mechanism and adds to earlier proposals for various histidine kinase architectures on scissoring piston-like movements, or rotation of the helical bundles 21,23 . Several mechanisms of signal transduction into the effector domains may coexist 25 , reflecting that histidine kinases integrate many different signals in a variety of architectures and have evolved through domain swapping 45 . Similarly, the photosensory modules of phytochromes may adapt their structural changes to the output domains, which may explain why they can form a variety of domain fusions with distinct effectors 16,17,40,46 .…”

“…A highly pertinent question is how the kinases achieve this signal transduction at the atomic level. Crystal structures of proteins involved in TCS have resulted in a number of partially conflicting conformational mechanisms for signal transduction [21][22][23][24][25][26] , one of which is that the region around the active site for phosphate transfer is destabilized 27 . Opposing the notion that one…”

“…clearly defined cascade of structural changes leads to signaling, it has been proposed that allosteric coupling between the different modules controls the activity of the effector domains 25,28 , but a structural understanding of this concept is missing.…”

Engineering transmembrane signal transduction in synthetic membranes using two-component systems

“…However, luminescence for the vancomycin sensor was highest across all conditions (Fig. 4E), suggesting that the chimera may be locked into an “on” conformation, perhaps due to destabilizing the HAMP domain ( 44 ). These results demonstrate how protein and membrane engineering can be used to readily generate new cell-free receptors by tapping into the large repertoire of known sensing domains of transmembrane proteins, and underscores the important relationship between membrane physical properties, protein sequence, and function.…”

“…3C). This nitrate-independent increase in luminescence upon addition of POPE may suggest that negative lipid curvature stabilizes the "on" conformation of NarX (44). The high induction of luciferase expression in POPE and POPG membranes is unsurprising as PE and PG make up the majority of lipid headgroups in E. coli membranes (45).…”