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A new approach to producing functional G alpha subunits yields the activated and deactivated structures of G alpha(12/13) proteins.
- Source :
-
Biochemistry [Biochemistry] 2006 Jan 10; Vol. 45 (1), pp. 167-74. - Publication Year :
- 2006
-
Abstract
- The oncogenic G(12/13) subfamily of heterotrimeric G proteins transduces extracellular signals that regulate the actin cytoskeleton, cell cycle progression, and gene transcription. Previously, structural analyses of fully functional G alpha(12/13) subunits have been hindered by insufficient amounts of homogeneous, functional protein. Herein, we report that substitution of the N-terminal helix of G alpha(i1) for the corresponding region of G alpha12 or G alpha13 generated soluble chimeric subunits (G alpha(i/12) and G alpha(i/13)) that could be purified in sufficient amounts for crystallographic studies. Each chimera bound guanine nucleotides, G betagamma subunits, and effector proteins and exhibited GAP responses to p115RhoGEF and leukemia-associated RhoGEF. Like their wild-type counterparts, G alpha(i/13), but not G alpha(i/12), stimulated the activity of p115RhoGEF. Crystal structures of the G alpha(i/12) x GDP x AlF4(-) and G alpha(i/13) x GDP complexes were determined using diffraction data extending to 2.9 and 2.0 A, respectively. These structures reveal not only the native structural features of G alpha12 and G alpha13 subunits, which are expected to be important for their interactions with GPCRs and effectors such as G alpha-regulated RhoGEFs, but also novel conformational changes that are likely coupled to GTP hydrolysis in the G alpha(12/13) class of heterotrimeric G proteins.
- Subjects :
- Animals
Binding Sites
Cells, Cultured
Chimera genetics
Chimera metabolism
Crystallography, X-Ray
GTP-Binding Protein alpha Subunits, G12-G13 genetics
GTP-Binding Protein alpha Subunits, G12-G13 isolation & purification
GTP-Binding Protein beta Subunits chemistry
GTP-Binding Protein beta Subunits genetics
GTP-Binding Protein beta Subunits metabolism
GTP-Binding Protein gamma Subunits chemistry
GTP-Binding Protein gamma Subunits genetics
GTP-Binding Protein gamma Subunits metabolism
GTPase-Activating Proteins chemistry
GTPase-Activating Proteins genetics
GTPase-Activating Proteins metabolism
Guanine Nucleotide Exchange Factors genetics
Guanine Nucleotide Exchange Factors metabolism
Guanine Nucleotides chemistry
Guanine Nucleotides genetics
Guanine Nucleotides metabolism
Leukemia metabolism
Protein Conformation
Proto-Oncogene Proteins chemistry
Proto-Oncogene Proteins genetics
Proto-Oncogene Proteins metabolism
Receptors, G-Protein-Coupled genetics
Receptors, G-Protein-Coupled metabolism
Rho Guanine Nucleotide Exchange Factors
GTP-Binding Protein alpha Subunits, G12-G13 chemistry
GTP-Binding Protein alpha Subunits, G12-G13 metabolism
Subjects
Details
- Language :
- English
- ISSN :
- 0006-2960
- Volume :
- 45
- Issue :
- 1
- Database :
- MEDLINE
- Journal :
- Biochemistry
- Publication Type :
- Academic Journal
- Accession number :
- 16388592
- Full Text :
- https://doi.org/10.1021/bi051729t