The crystal structure of the periplasmic domain of Vibrio parahaemolyticus CpxA

Abstract

The Cpx two‐component system of Gram‐negative bacteria senses extracytoplasmic stresses using the histidine kinase CpxA, a membrane‐bound sensor, and controls the transcription of the genes involved in stress response by the cytosolic response regulator CpxR, which is activated by the phosphorelay from CpxA. CpxP, a CpxA‐associated protein, also plays an important role in the regulation of the Cpx system by inhibiting the autophosphorylation of CpxA. Although the stress signals and physiological roles of the Cpx system have been extensively studied, the lack of structural information has limited the understanding of the detailed mechanism of ligand binding and regulation of CpxA. In this study, we solved the crystal structure of the periplasmic domain of Vibrio parahaemolyticus CpxA (VpCpxA‐peri) to a resolution of 2.1 Å and investigated its interaction with CpxP. VpCpxA‐peri has a globular Per‐ARNT‐SIM (PAS) domain and a protruded C‐terminal tail, which may be required for ligand sensing and CpxP binding, respectively. The direct interaction of the PAS core of VpCpxA‐peri with VpCpxP was not detected by NMR, suggesting that the C‐terminal tail or other factors, such as the membrane environment, are necessary for the binding of CpxA to CpxP.

Dynamic Interaction between the CpxA Sensor Kinase and the Periplasmic Accessory Protein CpxP Mediates Signal Recognition in E. coli

“…To control that cytosolic expression of the fusion proteins did not confer with correct folding we first confirmed recent crystallographic data showing that CpxP and CpxA-SD form homodimers (Fig. S1) [27]–[29]. Next, the cyclase-deficient E. coli strain BTH101 was co-transformed with pairs of fusion plasmids expressing either CpxP or CpxA-SD and the transformants were tested for their ability to metabolize lactose on MacConkey agar plates (Fig.…”

“…Accomplishing peptide arrays indicate that the C-terminal region of the periplasmic sensor domain of CpxA (E 138 DNYOLYLIRPASSSQSDEINLLFD 162 ) might play an important role for interaction with CpxP [27]. However, NMR studies could not detect a direct interaction between the periplasmic domain of Vibrio parahaemolyticus CpxA (VpCpxA-peri) and the respective CpxP protein (VpCpxP) [29].…”

Envelope Stress Responses: An Interconnected Safety Net

“…A recent structure of the Vibrio parahaemolyticus CpxA sensory domain shows it forms a PAS domain of five β-strands and three α-helices [37]. Mutations in the sensory domain can activate CpxA.…”

“…Mutations in the sensory domain can activate CpxA. For example, the well-studied cpxA24 mutation deletes 32 residues and entirely removes a C-terminal sensory domain α-helix [36,37]. CpxA24 is constitutively activated and signal blind, suggesting that disrupting proper sensor domain folding may directly trigger CpxA kinase activity [36,38].…”

Prokaryotic 2-component systems and the OmpR/PhoB superfamily

“…Auxiliary proteins that interact with HKs can turn on or off the signal transduction of a TCS. An example is the E. coli periplasmic protein CpxP, which interacts with the periplasmic domain of the sensor CpxA and turns off the CpxA-CpxR TCS (Fleischer et al 2007;Buelow and Raivio 2010;Kwon et al 2012;Debnath et al 2013). These additional mechanisms for fine regulation can also be viewed as ways to accommodate changes in the circumstances of a singlecelled organism.…”