Abstract
ABSTRACTRas GTPase is a peripheral membrane protein central to cellular signaling of growth and proliferation. Membrane attachment is critical for a range of Ras activities, therefore, ability to make faithful in-vitro samples of a mem-brane-bound Ras for detailed biophysical studies is a highly desirable goal. In this manuscript, we are describing preparation of a large-scale sample of isotopically labeled H-Ras conjugated to lipid nanodiscs. We demonstrate that the Ras-nanodisc sample is fairly stable to allow for a range of Nuclear Magnetic Resonance (NMR) and other biophysical measurements. The need to achieve a homogeneous protein-nanodisc ratio is also emphasized.
Publisher
Cold Spring Harbor Laboratory
Reference27 articles.
1. Colicelli, J. (2004) Human RAS Superfamily Proteins and Related GTPases, Sci. STKE 2004, re13.
2. Wittinghofer, A. , and Vetter, I. R. (2011) Structure-Function Relationships of the G Domain, a CanonicalSwitch Motif, In Annual Review of Biochemistry ( Kornberg, R. D. , Raetz, C. R. H. , Rothman, J. E. , and Thorner, J. W. , Eds.), pp 943’971, Annual Reviews, Palo Alto.
3. All ras proteins are polyisoprenylated but only some are palmitoylated
4. Ras trafficking, localization and compartmentalized signalling
5. Ras–effector interactions: after one decade