Flexibility of the Cytoplasmic Domain of the Phototaxis Transducer II from Natronomonas pharaonis

Abstract

Chemo- and phototaxis systems in bacteria and archaea serve as models for more complex signal transduction mechanisms in higher eukaryotes. Previous studies of the cytoplasmic fragment of the phototaxis transducer (pHtrII-cyt) from the halophilic archaeon Natronomonas pharaonis showed that it takes the shape of a monomeric or dimeric rod under low or high salt conditions, respectively. CD spectra revealed only approximately 24% helical structure, even in 4 M KCl, leaving it an open question how the rod-like shape is achieved. Here, we conducted CD, FTIR, and NMR spectroscopic studies under different conditions to address this question. We provide evidence that pHtrII-cyt is highly dynamic with strong helical propensity, which allows it to change from monomeric to dimeric helical coiled-coil states without undergoing dramatic shape changes. A statistical analysis of predicted disorder for homologous sequences suggests that structural flexibility is evolutionarily conserved within the methyl-accepting chemotaxis protein family.

Molecular model of a sensor of two-component signaling system

“…In the latter, the very long cytoplasmic domain of the dimeric Np HtrII transducer appear to be highly flexible, corroborating the intrinsically dynamic nature of chemo- and photoreceptors described in some recent publications (e.g. 10 , 40 , 44 ). Coupled with the conclusions of a previous study that the cytoplasmic domain of Np HtrII's ( p HtrII-cyt) does not form dimers at low ionic strength 33 , our results imply that only the transmembrane region is required for dimerization of Np SRII/ Np HtrII.…”

“…Nevertheless, we cannot rule out that such conformational flexibility may be a result of low ionic strength in which the transducer domain is partially disordered. Indeed, in line with the results of other studies 33 , 40 , CD-spectra of the full-length Np SRII/ Np HtrII complex under different salt conditions (Fig. S1 ) suggest partial disordering of Np SRII/ Np HtrII dimers.…”

“…Due to the difficulty to unequivocally determine the amount of random coil structures, these authors argued in light of their SANS in favor a highly dynamic helical rod at low salt conditions. This finding has been confirmed by subsequent results published in the work 40 , according to which p HtrII-cyt has a strong propensity for helix. Firstly, p HtrII-cyt has a high helix propensity inferred from the sequence; p HtrII-cyt is predicted to be a helical coiled-coil based on a homology to the Tst-cyt 22 .…”

“…Thus, despite "random-coil" spectroscopic features, p HtrII-cyt can exist as a highly flexible, loosely packed but folded helical coiled-coil. The results of the comparison of CD spectra from the work 40 with and without TFE suggest that, despite the presence of β-sheets in the deconvolution of CD spectra (see Fig. S1 B), β-sheets do not necessarily have to be present in the final molecular model.…”

“…2 A). In order to improve the agreement between theoretical and experimental scattering curves data, we generated models of different possible conformations of the Np SRII/ Np HtrII dimers which, while maintaining an elongated shape for the Np HtrII dimers, considered both bending at its flexible hinges 11 and partial unfolding 33 , 40 of this domain at low salt conditions. Our results confirm a highly dynamic nature of the transducer dimer at low salt: the root mean square deviations of bending angles from the zero value are in the range from 50° to 60° for all the three HAMP1-, HAMP2-, and Gly- hinges (see Fig.…”