Motile microorganisms navigate through their environment using special molecular machinery in order to sense gradients of various signals: chemotaxis (reactions to chemical compounds) and phototaxis (to light) sensory cascades. Transmembrane receptors play a central role in these cascades as they receive input signals and transmit them inside the cell, where they modulate activity of the kinases CheA, which are tightly bound to their cytoplasmic domains. CheA further phosphorylates the response regulator protein CheY, which regulates the flagella. At the same time, CheA phosphorylates and, by means of this, activates another response regulator, CheB, which, along with the constantly active CheR protein, catalyzes two opposite reactions: methylation and demethylation of the specific glutamic acid residues located at the cytoplasmic domain of the receptors. The latter reactions establish the adaptation mechanism, which allows microbes to sense in a very broad range of the input signal intensities.
Many functional, structural and dynamical aspects of the signal propagation through the prokaryotic receptors as well as a mechanism of the signal amplification remain still unclear. In the present thesis we have used various techniques of computational biophysics, chiefly molecular dynamics (MD) simulations, in order to approach these problems.
In Chapter 3, we have carried out MD simulations of the isolated linker domain (HAMP) from the E. coli Tsr chemoreceptor. The MD simulations revealed highly dynamical nature of this domain, which allows for interconversion between several metastable states. These metastable states feature a number of structural and dynamical properties, which were previously reported for HAMP domains of various receptors obtained from different organisms. It allowed us to reconcile numerous experimental data and to hypothesize that different HAMP domains share similar mechanism of their action.
In Chapter 4, we have performed MD simulations of the whole cytoplasmic domain of the Tsr chemoreceptor. The simulations revealed a mechanism for the inter-domain coupling between the HAMP domain and a part of the cytoplasmic domain adjacent to the HAMP, the adaptation subdomain, by means of the regulated unfolding of a short linker region termed the stutter. Also, we have found that the reversible methylation/demethylation of the cytoplasmic domain affects its flexibility and symmetry. Altogether, these findings suggest a mechanism of signal propagation at the level of an individual chemoreceptor dimer.
In Chapter 5, we have built a model of the trimer-of-dimers of the archaeal phototaxis receptor complex (NpSRII:NpHtrII). Subsequent MD simulations revealed an important role of dynamics in signal transduction and, potentially, in the kinase activation.
In Chapter 6, we have reconstructed a whole transmembrane lattice formed by the NpSRII:NpHtrII complexes. The concave shape of the obtained lattice naturally explains polar localization of the receptor arrays in prokaryotic cells. At the same time, additional MD simulations of an individual unit of this lattice (a dimer of the photosensor) revealed global motional modes in its transmembrane region, which presumably co-occur with its activation and can spread across the tightly packed transmembrane arrays allowing for the signal amplification.
| Identifer | oai:union.ndltd.org:uni-osnabrueck.de/oai:repositorium.ub.uni-osnabrueck.de:urn:nbn:de:gbv:700-2016081014821 |
| Date | 10 August 2016 |
| Creators | Orekhov, Philipp S |
| Contributors | Prof. Dr. Heinz-Jürgen Steinhoff, Prof. Dr. Konstantin V. Shaitan |
| Source Sets | Universität Osnabrück |
| Language | English |
| Detected Language | English |
| Type | doc-type:doctoralThesis |
| Format | application/zip, application/pdf |
| Rights | http://rightsstatements.org/vocab/InC/1.0/ |
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