Mode Of Action Of The Bacterial Thermosensor Desk Involved In Regulating Membrane Fluidity

The Bacillus subtilis membrane harbors the temperature sensing and signaling protein DesK. At low temperatures it triggers expression of a desaturase, which introduces double bonds into pre-existing phospholipids, thereby regulating membrane fluidity. Recently it was discovered [1] that both sensing and transmission of DesK, which has five transmembrane segments, can be captured into one single chimerical transmembrane segment, the so-called ‘minimal sensor' (DesK-MS). This protein can be functionally reconstituted in lipid bilayers, thus providing an excellent model system to study the molecular details of a biologically important signaling mechanism. Here we used synthetic peptides corresponding to functional and non-functional mutants of the minimal sensor in artificial membranes of phosphatidylcholines as convenient model systems. We studied the conformational properties, tilt, and exposure at the lipid/water interface at different bilayer thickness and upon varying temperature by using circular dichroism and fluorescence studies. Based on these results and on mutational studies, we propose a model for the mode of action of DesK-MS, in which an N-terminal “sunken buoy” motif and a C-terminal hydrophilic motif are crucial for DesK-MS functioning. Finally, we explored the possibility of isolating and characterizing DesK-MS in its native membrane in the form of “native nanodiscs” by using copolymers of styrene and maleic acid (SMA). Results of these studies will be presented. [1] Cybulski LE, Martin M, Mansilla MC, Fernandez A, de Mendoza D. Membrane thickness cue for cold sensing in a bacterium. Curr Biol. 2010 20(17):1539-44