[NMR paper] Membrane-dependent modulation of the mTOR activator Rheb: NMR observations of a GTPase tethered to a lipid-bilayer nanodisc.

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Membrane-dependent modulation of the mTOR activator Rheb: NMR observations of a GTPase tethered to a lipid-bilayer nanodisc.

Membrane-dependent modulation of the mTOR activator Rheb: NMR observations of a GTPase tethered to a lipid-bilayer nanodisc.

J Am Chem Soc. 2013 Feb 14;

Authors: Mazhab-Jafari MT, Marshall CB, Stathopulos PB, Kobashigawa Y, Stambolic V, Kay LE, Inagaki F, Ikura M

Abstract
Like most Ras superfamily proteins, the GTPase domain of Ras homolog enriched in brain (Rheb) is tethered to cellular membranes through a prenylated cysteine in a flexible C-terminal region, however little is known about how Rheb or other GTPases interact with the membrane or how this environment may affect their GTPase functions. We used NMR methods to characterize Rheb tethered to nanodiscs, monodisperse protein-encapsulated lipid bilayers with a diameter of 10 nm. Membrane conjugation markedly reduced the rate of intrinsic nucleotide exchange, while GTP hydrolysis was unchanged. NMR measurements revealed that the GTPase domain interacts transiently with the surface of the bilayer with two distinct preferred orientations, which are determined by the bound nucleotide. We propose models of membrane-dependent signal regulation by Rheb that shed light on previously unexplained in vivo properties of this GTPase. The study presented provides a general approach for direct ex-perimental investigation of membrane-dependent properties of other Ras-superfamily GTPases.


PMID: 23409921 [PubMed - as supplied by publisher]



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