Determination of structural requirements for the interaction of Rab6 with RabGDI and Rab geranylgeranyltransferase.
The importance of geranylgeranylation to the interaction of Rab proteins with RabGDI was investigated with a set of Rab6 mutants post-translationally modified by all known C-terminal lipid combinations. Rab6 proteins geranylgeranylated on CXC or CC motifs were found to be significantly better substrates for membrane extraction by RabGDI than either Rab6 proteins geranylgeranylated on CAAL motifs or Rab6 proteins that were farnesylated and palmitoylated. The methylation status of the CXC motif did not significantly affect interaction of wild type Rab6 with RabGDI. Rab6 protein sequences required for RabGDI interaction were then identified. Consistent with the significant homology between Rab-GDI and the Rab escort protein, a subunit of Rab geranylgeranyltransferase (RabGGTase), we show that there is an overlap between Rab6 motifs required for RabGDI binding and RabGGTase processing. The effector domain, loop3/beta 3 and the hypervariable region of Rab6 are all required for RabGDI binding, whereas loop3/beta 3 and the hypervariable region but not the effector domain are required for efficient processing of Rab6 by RabGGTase. Interestingly, however, loop3/beta 3 of Rab6 when introduced into H-Ras is sufficient to allow some in vivo processing of a C-terminal CSC motif.
Beranger
F
F
INSERM Unite 248, Paris.
Cadwallader
K
K
Porfiri
E
E
Powers
S
S
Evans
T
T
de Gunzburg
J
J
Hancock
J F
JF
D019883:Alkyl and Aryl Transferases; D000595:Amino Acid Sequence; D002478:Cells, Cultured; D019204:GTP-Binding Proteins / Q000737:chemistry / Q000378:metabolism*; D020730:Guanine Nucleotide Dissociation Inhibitors; D006801:Humans; D008969:Molecular Sequence Data; D011499:Protein Processing, Post-Translational; D014166:Transferases / Q000378:metabolism*
C086796:GDP dissociation inhibitor 1; D020730:Guanine Nucleotide Dissociation Inhibitors; D014166:Transferases; D019883:Alkyl and Aryl Transferases; C075607:Rab geranylgeranyltransferase; D019204:GTP-Binding Proteins