Clustering and dynamics of phototransducer signaling domains revealed by SDSL EPR on SRII/HtrII in membranes and nanodiscs.

Biochemistry

PubMedID: 25489970

Orban-Glaß I, Voskoboynikova N, Busch KB, Klose D, Rickert C, Mosslehy W, Roder F, Wilkens V, Piehler J, Engelhard M, Steinhoff H, Klare JP. Clustering and dynamics of phototransducer signaling domains revealed by SDSL EPR on SRII/HtrII in membranes and nanodiscs. Biochemistry. 2014;.
In halophilic archaea the photophobic response is mediated by the membrane-embedded 2:2 photoreceptor/-transducer complex SRII/HtrII, the latter being homologous to the bacterial chemoreceptors. Both systems bias the rotation direction of the flagellar motor via a two-component system coupled to an extended cytoplasmic signaling domain formed by a four helical anti-parallel coiled-coil structure. For signal propagation by the HAMP domains connecting the transmembrane and cytoplasmic domains it was suggested that a two-state thermodynamic equilibrium found for the first HAMP domain in NpSRII/NpHtrII is shifted upon activation, yet signal propagation along the coiled-coil transducer remains largely elusive, including the activation mechanism of the coupled kinase CheA. We investigated the dynamic and structural properties of the cytoplasmic tip domain of NpHtrII in terms of signal transduction and putative oligomerization using site-directed spin labeling (SDSL) electron paramagnetic resonance (EPR) spectroscopy. We show that the cytoplasmic tip domain of NpHtrII is engaged in a two-state equilibrium between a dynamic and a compact conformation like it was found for the first HAMP domain, thus strengthening the assumption that dynamics are the language of signal transfer. Inter spin distance measurements in membranes and on isolated 2:2 photoreceptor/transducer complexes in nanolipoprotein particles (NLPs) provide evidence that archaeal photoreceptor/-transducer complexes analogous to chemoreceptors form trimers-of-dimers or higher-order assemblies even in the absence of the cytoplasmic components CheA and CheW, underlining conservation of the overall mechanistic principles underlying archaeal phototaxis and bacterial chemotaxis systems. Furthermore, our results revealed a significant influence of the NpHtrII signaling domain on the NpSRII photocycle kinetics, providing evidence for a conformational coupling of SRII and HtrII in these complexes.