ИСТИНА

P75NTR is a type I integral membrane protein, which plays a key role in neurotrophin signaling. The structural data, describing the receptor in various functional states is sparse and controversial. The transmembrane domain (TM) domain of p75NTR plays a key role in receptor function. This domain stabilizes receptor dimers through a disulfide bond essential for p75NTR. Using NMR spectroscopy we solved the three-dimensional structure of p75-TM in lipid micelles. We describe two different dimer conformations: the wild-type forms a covalent dimer stabilized by a disulfide bond, while the C257A mutant forms a non-covalent dimer via an interface on the opposite side of the a-helix. These p75 dimers differ in terms of crossing angle and γ- secretase cleavage. We additionally investigated the connection between the states of TMD and intracellular domain (ICD) of p75NTR, in order to get an insight into the details of the molecular activation mechanism of the receptor. For that purpose, we used two constructs on the basis of the p75NTR: (1) construct, containing TMD and juxtamembrane “chopper” domain, and (2) construct, containing TMD and full-size ICD of the receptor. These constructs were incorporated into various membrane mimetics, including lipid/protein nanodiscs of different size and composition. We managed to obtain NMR spectra of constructs with folded “Death Domain” (DD), assign chemical shifts and study the intramolecular mobility of p75NTR ICD or “chopper” domain in lipid/protein nanodiscs and in almost physiological conditions. Our 608 data reveal that the whole ICD is extremely mobile and partially disordered, which results in a loose linkage between the motions of the ICD and motions of the TMD helix. Moreover, our experiments with lipid/protein nanodiscs, containing several copies of constructs on the basis of p75NTR, demonstrate that DDs of the receptor have no propensity to interact tightly with each other or to bind to the membrane. As a result, previously suggested activation mechanism of p75NTR is brought into question with our observations. With this respect, we discuss our data in the context of the receptor activation and suggest two possible mechanisms that contradict neither the reported NMR data, nor the previously published results of functional assays and other in vivo studies. This work was supported by the Russian Foundation for Basic Research (project No. 13-04-40107 сomfi).