Kremer, Werner and Steiner, Guido and Beraud-Dufour, Sophie and Kalbitzer, Hans Robert (2004) Conformational states of the small G protein Arf-1 in complex with the guanine nucleotide exchange factor ARNO-Sec7. JOURNAL OF BIOLOGICAL CHEMISTRY, 279 (17). pp. 17004-17012. ISSN 0021-9258, 1083-351X
Full text not available from this repository. (Request a copy)Abstract
Arfl is a small G protein involved in vesicular trafficking, and although it is only distantly related to Ras, it adopts a similar three-dimensional structure. In the present work, we study Arfl bound to GDP and GTP and its interactions with one of its guanosine nucleotide exchange factors, ARNO-Sec7. The P-31 NMR spectra of Arfl.GDP.Mg2+ and Arfl.GTP.Mg2+ share the general features typical for all small G proteins studied so far. Especially, the beta-phosphate resonances of the bound nucleotide are shifted strongly downfield compared with the resonance positions of the free magnesium complexes of GDP and GTP. However, no evidence for an equilibrium between two conformational states of Arfl.GDP.Mg-2 or Arfl.GTP.Mg2+ could be observed as it was described earlier for Ras and Ran. Glu(156) of ARNO-Sec7 has been suggested to play as "glutamic acid finger" an important role in the nucleotide exchange mechanism. In the millimolar concentration range used in the NMR experiments, wild type ARNO-Sec7 and ARNO-Sec7(E156D) do weakly interact with Arfl.GDP.Mg2+ but do not form a strong complex with magnesium-free Arfl.GDP. Only wild type ARNO-Sec7 competes weakly with GDP on Arfl.GDP.Mg2+ and leads to a release of GDP when added to the solution. The catalytically inactive mutants ARNO-Sec7(E156A) and ARNO-Sec7(E156K) induce a release of magnesium from Arfl.GDP.Mg2+ but do not promote GDP release. In addition, ARNO-Sec7 does not interact or only very weakly interacts with the GTP-bound form of Arf1, opposite to the observation made earlier for Ran, where the nucleotide exchange factor RCC1 forms a complex with Ran.GTP.Mg2+ and is able to displace the bound GTP.
| Item Type: | Article |
|---|---|
| Uncontrolled Keywords: | FACTOR ARNO; BINDING; SEC7; GDP; P-31; PHOSPHOLIPIDS; ACTIVATION; DOMAINS; FAMILY; GTPASE; |
| Subjects: | 500 Science > 570 Life sciences |
| Divisions: | Biology, Preclinical Medicine > Institut für Biophysik und physikalische Biochemie > Prof. Dr. Dr. Hans Robert Kalbitzer |
| Depositing User: | Dr. Gernot Deinzer |
| Date Deposited: | 21 Jul 2021 08:11 |
| Last Modified: | 21 Jul 2021 08:11 |
| URI: | https://pred.uni-regensburg.de/id/eprint/37736 |
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