Structural basis for the association of the redox-sensitive target of rapamycin FATC domain with membrane-mimetic micelles.
The target of rapamycin (TOR) is a conserved eukaryotic Ser/Thr kinase that regulates cellular growth in response to the nutrient and energy state. TOR signaling plays an important role in the development of diseases such as cancer, obesity, and diabetes and in different redox-sensitive processes (hypoxia, apoptosis, and aging). Because TOR has been detected at different cellular membranes and in the nucleus, its localization may influence the specific signaling readout. To better understand how TOR can associate with different membranes, the lipid-binding properties of the redox-sensitive yeast TOR1 FATC domain (y1fatc) have been characterized by solution NMR spectroscopy. Binding studies with different lipids indicate that y1fatc interacts specifically with a membrane-mimetic environment but appears not to recognize a specific lipid headgroup. In both, the structures of oxidized and reduced micelle-bound y1fatc, residues Ile-2456 to Trp-2470 of the lipid-binding motif form a hydrophobic bulb that has a rim of charged residues. The diffusion constants for both micelle-bound states are consistent with the rotational correlation times from the analysis of the (15)N relaxation data. Based on the K(d) values, the oxidized form (K(d) approximately 0.31 mm) binds dodecyl phosphocholine micelles slightly tighter than the reduced form (K(d) approximately 1.86 mM). Binding studies with y1fatc in which one or both tryptophans (Trp-2466 and Trp-2470) were replaced by alanine suggest that these residues are important for the exact positioning in the membrane and that the other aromatic (His-2462, Tyr-2463, and Phe-2469) and aliphatic residues (Ile-2456, Leu-2459, Ile-2464, and Pro-2468) in the lipid-binding motif contribute significantly to the affinity.
Dames
Sonja A
SA
Department of Structural Biology, Biozentrum, University of Basel, Klingelbergstrasse 70, 4056 Basel, Switzerland. sonja.dames@tum.de
D000595:Amino Acid Sequence; D008563:Membrane Lipids / Q000737:chemistry / Q000378:metabolism; D008823:Micelles; D008958:Models, Molecular; D008969:Molecular Sequence Data; D019906:Nuclear Magnetic Resonance, Biomolecular; D010084:Oxidation-Reduction; D019869:Phosphatidylinositol 3-Kinases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D011485:Protein Binding; D011487:Protein Conformation; D017434:Protein Structure, Tertiary; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D016415:Sequence Alignment
D008563:Membrane Lipids; D008823:Micelles; D029701:Saccharomyces cerevisiae Proteins; D019869:Phosphatidylinositol 3-Kinases; C083324:TOR1 protein, S cerevisiae