C-terminal binding domain of Rho GDP-dissociation inhibitor directs N-terminal inhibitory peptide to GTPases.
The Rho GDP-dissociation inhibitors (GDIs) negatively regulate Rho-family GTPases. The inhibitory activity of GDI derives both from an ability to bind the carboxy-terminal isoprene of Rho family members and extract them from membranes, and from inhibition of GTPase cycling between the GTP- and GDP-bound states. Here we demonstrate that these binding and inhibitory functions of rhoGDI can be attributed to two structurally distinct regions of the protein. A carboxy-terminal folded domain of relative molecular mass 16,000 (M[r] 16K) binds strongly to the Rho-family member Cdc42, yet has little effect on the rate of nucleotide dissociation from the GTPase. The solution structure of this domain shows a beta-sandwich motif with a narrow hydrophobic cleft that binds isoprenes, and an exposed surface that interacts with the protein portion of Cdc42. The amino-terminal region of rhoGDI is unstructured in the absence of target and contributes little to binding, but is necessary to inhibit nucleotide dissociation from Cdc42. These results lead to a model of rhoGDI function in which the carboxy-terminal binding domain targets the amino-terminal inhibitory region to GTPases, resulting in membrane extraction and inhibition of nucleotide cycling.
Gosser
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YQ
Cellular Biochemistry and Biophysics Program, Memorial Sloan-Kettering Cancer Center, New York 10021, USA.
Nomanbhoy
T K
TK
Aghazadeh
B
B
Manor
D
D
Combs
C
C
Cerione
R A
RA
Rosen
M K
MK
D000595:Amino Acid Sequence; D000818:Animals; D002417:Cattle; D018797:Cell Cycle Proteins / Q000737:chemistry / Q000378:metabolism*; D020558:GTP Phosphohydrolases / Q000378:metabolism*; D019204:GTP-Binding Proteins / Q000737:chemistry* / Q000378:metabolism*; D020730:Guanine Nucleotide Dissociation Inhibitors; D006153:Guanosine Diphosphate / Q000031:analogs & derivatives / Q000378:metabolism*; D006801:Humans; D006860:Hydrogen Bonding; D009682:Magnetic Resonance Spectroscopy; D008958:Models, Molecular; D008969:Molecular Sequence Data; D008970:Molecular Weight; D011485:Protein Binding; D011487:Protein Conformation; D017510:Protein Folding; D017368:Protein Prenylation; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D016415:Sequence Alignment; D013816:Thermodynamics; D020846:cdc42 GTP-Binding Protein, Saccharomyces cerevisiae; D062367:ortho-Aminobenzoates / Q000378:metabolism; D062948:rho Guanine Nucleotide Dissociation Inhibitor alpha; D062947:rho-Specific Guanine Nucleotide Dissociation Inhibitors
C569638:ARHGDIA protein, human; D018797:Cell Cycle Proteins; D020730:Guanine Nucleotide Dissociation Inhibitors; D062367:ortho-Aminobenzoates; D062948:rho Guanine Nucleotide Dissociation Inhibitor alpha; D062947:rho-Specific Guanine Nucleotide Dissociation Inhibitors; C065197:3'-(methylanthraniloyl)-2'-deoxy-guanosine diphosphate; D006153:Guanosine Diphosphate; D020558:GTP Phosphohydrolases; D019204:GTP-Binding Proteins; D020846:cdc42 GTP-Binding Protein, Saccharomyces cerevisiae