Characterization of the Intrinsic and TSC2-GAP–Regulated GTPase Activity of Rheb by Real-Time NMR

Christopher B. Marshall,^1,2 Jason Ho,¹ Claudia Buerger,^1,2 Michael J. Plevin,^1,2 Guang-Yao Li,^1,2 Zhihong Li,^1,2 Mitsuhiko Ikura,^1,2* Vuk Stambolic^1,2*

This PDF file includes:

• Methods
• Fig. S1. Role of Rheb in mTOR signaling.
• Fig. S2. Spectra of Rheb-GDP and Rheb-GMPPNP.
• Fig. S3. Secondary structure and chemical shift index analysis.
• Fig. S4. Preparation of Rheb-GTP for GTPase assay.
• Fig. S5. Time course of GTP hydrolysis of Rheb-GTP.
• Fig. S6. Analysis of Rheb-GTP and Rheb-GDP during GTP hydrolysis.
• Fig. S7. Circular dichroism spectra of WT TSC2_1225–1742 and its variants.
• Fig. S8. Further development of real-time NMR method to increase temporal
• resolution.
• Fig. S9. Further development of real-time NMR method to decrease sample requirement.
• References

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¹Division of Signaling Biology, Ontario Cancer Institute, University Health Network, Toronto, Ontario, Canada M5G 2M9. ²Department of Medical Biophysics, University of Toronto, Toronto, Ontario, Canada M5G 2M9.

*To whom correspondence should be addressed. E-mail, vuks{at}uhnres.utoronto.ca (V.S.) and mikura{at}uhnresearch.ca (M.I.)

Citation: C. B. Marshall, J. Ho, C. Buerger, M. J. Plevin, G.-Y. Li, Z. Li, M. Ikura, V. Stambolic, Characterization of the Intrinsic and TSC2-GAP–Regulated GTPase Activity of Rheb by Real-Time NMR. Sci. Signal. 2, ra3 (2009).

© 2009 American Association for the Advancement of Science