Signaling and Adaptation Modulate the Dynamics of the Photosensoric Complex of Natronomonas pharaonis

Abstract

Motile bacteria and archaea respond to chemical and physical stimuli seeking optimal conditions for survival. To this end transmembrane chemo- and photoreceptors organized in large arrays initiate signaling cascades and ultimately regulate the rotation of flagellar motors. To unravel the molecular mechanism of signaling in an archaeal phototaxis complex we performed coarse-grained molecular dynamics simulations of a trimer of receptor/transducer dimers, namely NpSRII/NpHtrII from Natronomonas pharaonis. Signaling is regulated by a reversible methylation mechanism called adaptation, which also influences the level of basal receptor activation. Mimicking two extreme methylation states in our simulations we found conformational changes for the transmembrane region of NpSRII/NpHtrII which resemble experimentally observed light-induced changes. Further downstream in the cytoplasmic domain of the transducer the signal propagates via distinct changes in the dynamics of HAMP1, HAMP2, the adaptation domain and the binding region for the kinase CheA, where conformational rearrangements were found to be subtle. Overall these observations suggest a signaling mechanism based on dynamic allostery resembling models previously proposed for E. coli chemoreceptors, indicating similar properties of signal transduction for archaeal photoreceptors and bacterial chemoreceptors.

Fig 1. Two component phototaxis system of N. pharaonis.

NpSRII/NpHtrII dimers are the basic elements of photoreceptor complexes in N. pharaonis. They consist of two sensory rhodopsins, NpSRII, and two transducer proteins, NpHtrII, mostly of α-helical secondary structure, with a characteristic domain organization. Light activation of NpSRII induces conformational and/or dynamical changes in the transducer which are converted by two HAMP domains and conveyed along the 20 nm long transducer to the tip region, where it activates the homodimeric histidine kinase CheA bound together with the adapter protein CheW. The kinase CheA undergoes auto-phosphorylation and further transfers the phosphate group to the response regulators CheY or CheB. CheY affects the rotational bias of the flagellar motor, while the methylesterase CheB along with the methyltransferase CheR controls the adaptation (feedback) mechanism. The related chemoreceptor and most likely also the photoreceptor dimers further organize into trimers, which, together with CheA and CheW, lead to the formation of large sensor arrays.

Fig 2. Conformations of the trimeric photoreceptor-transducer complexes.

Cartoon with the resulting structures of the demethylated (left) and the methylated (right) trimer systems combined in the bent membrane with a schematic representation of the adaptation process. Methylation sites are shown as red and black spheres in the demethylated and methylated state, respectively.

Fig 3. Intra-dimer distance changes between related residues of the transducer upon demethylation

A: Structure of the NpSRII/NpHtrII trimeric complex with distance changes color coded (calculated as an average over the three dimers). Positive values of the distance difference (blue) indicate a looser packing of the corresponding residues in the demethylated system, negative values (red) indicate a more compact packing. B: The intra-dimer distance difference as function of residue number shows distinct changes in the transmembrane region of the complex, an inversion of the packing densities for the two HAMP domains, minor changes at the methylation sites (m.s.), and a decrease of the packing density in the CheA/CheW binding site region labeled A/W.

Fig 4. Comparison of the conformational changes of the transmembrane region upon demethylation and illumination.

Cytoplasmic view of a monomer of the NpSRII/NpHtrII complex with experimentally observed conformational changes upon illumination shown as arrows and corresponding numerical values from the CG MD simulations. An outward tilt of helix F at the cytoplasmic side of the membrane embedded part of the transducer by 0.6±0.3 Å (blue arrow) is accompanied by a rotation of helix TM2 of 12±8° (olive green arrow) with respect to the equilibrated methylated structure. In addition TM2 shifts with respect to the helix TM1 of the transducer by 0.7±0.5 Å (light green arrow).

Fig 5. Inter-dimeric distances for related residues of the transducer.

Distances were calculated as an average over the three dimers for the methylated (black) and demethylated (red) states, shaded areas representing the standard deviation. The distance is measured between the center of mass (COM) of two related residues in one dimer and the COM of the six respective residues in the trimer-of-dimers (see inset on the lower left). The domains of the complex are depicted in colored bars; m.s and A/W indicate methylation sites and binding sites for CheA/CheW, respectively. Representative distance trajectories are depicted in S7 Fig.

Fig 6. Dynamics of the methylated and the demethylated systems.

A: Structure of the NpSRII/NpHtrII trimeric complex with colors that code for the difference between the RMSF value per residue of the demethylated and the methylated transducer. Positive values (in Å) correspond to a higher fluctuation and therefore higher mobility of the corresponding residues in the demethylated system, negative values indicate a lower mobility. B: The differences in mobility as function of residue number show distinct changes in the transmembrane region of the complex, an inversion between the two HAMP domains and in the adaptation and close to the glycine rich (293, 296) regions. This change in dynamics upon adaption includes the tip region and the binding sites for CheA (A/W). Colored bars have the same meaning as in Fig 3.

Fig 7. Model for the NpSRII/NpHtrII complex activation.

The regions with higher mobility are shown in diffuse representation; the arrows correspond to the domain motions (compacting/expanding within the trimer).