Purification and characterization from rat liver cytosol of a GDP dissociation inhibitor (GDI) for liver 24K G, a ras p21-like GTP-binding protein, with properties similar to those of smg p25A GDI

Abstract

A regulatory protein for a liver GTP-binding protein (G protein) with a molecular weight value of 24,000 (24K G), which we have recently purified, was purified to near-homogeneity from rat liver cytosol and characterized. This regulatory protein, designated here as GDP dissociation inhibitor for 24K G (24K G GDI), inhibited the dissociation of GDP from and the subsequent binding of GTP to 24K G. 24K G GDI was inactive for other ras p21/ras p21-like small G proteins including c-Ha-ras p21, rhoB p20, smg p21B, and smg p25A. 24K G was, however, recognized by bovine brain smg p25A GDI which regulated the GDP/GTP exchange reaction of smg p25A. By analyses of sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis (PAGE), immunoblotting with anti-smg p25A GDI antibody, two-dimensional PAGE, and C4 column chromatography, 24K G GDI showed physical properties very similar to those of smg p25A GDI. The peptide map and the partial amino acid sequences of 24K G GDI were not identical with those of smg p25A GDI. Among the 83 residues, 2 amino acids were different between rat liver 24K G GDI and bovine brain smg p25A GDI. These results indicate that there is a specific regulatory protein for 24K G, 24K G GDI, in rat liver cytosol and that 24K G GDI has close similarity to smg p25A GDI.

Interaction of Sec4 with GDI proteins from bovine brain, Drosophila melanogaster and Saccharomyces cerevisiae

“…Previ-236 ously it has been reported that rab GDI will inhibit dissociation of GDP from rab3A, rabll and Sec4 [26,28,29]. Rab GDI can also regulate the subcellular localization of rab3A by causing the dissociation of GDP-bound rab3A from synaptic membranes and vesicles [27].…”

“…By forming a stable soluble complex with prenylated, GDP-bound rab3A, rab3A GDI will prevent attachment of rab3A to membranes [27]. Rab3A GDI can also slow the dissociation of GDP from Sec4 [28] and rabll [29] and will regulate the membrane association of numerous rab proteins [30,31]. Because rab3A GDI has now been shown to be active on many members of the rab family, the protein has been renamed rab GDI [31].…”

“…Interestingly, a second GDI protein has been isolated which is closely related in sequence to rab GDI. This protein will inhibit dissociation of GDP from rabll but not from rab3A and has been given the name rabll GDI [29]. Neither rab GDI nor rabll GDI will interact with members of the ras or rho subgroups of the ras superfamily, suggesting that there maybe a family of sequence related GDI proteins specific for the rab proteins [27,29].…”

“…This protein will inhibit dissociation of GDP from rabll but not from rab3A and has been given the name rabll GDI [29]. Neither rab GDI nor rabll GDI will interact with members of the ras or rho subgroups of the ras superfamily, suggesting that there maybe a family of sequence related GDI proteins specific for the rab proteins [27,29]. Taken together, these observations suggest that rab GDI proteins may play an important role in the recycling of rab proteins from their target membranes through a soluble pool onto newly synthesized transport vesicles.…”

Starvation and ULK1-dependent cycling of mammalian Atg9 between the TGN and endosomes

“…The source of the AV membranes in mammalian cells is controversial: some studies (Dunn, Jr, 1990;Ericsson, 1969;Novikoff and Shin, 1978;Ueda et al, 1991) have identified endoplasmic reticulum (ER) proteins on the AVi, others detected Golgi membrane proteins (Locke and Sykes, 1975;Yamamoto et al, 1990). The unusual morphological properties of AVs has also led to the hypothesis that these membranes are derived from a unique type of membrane, called a phagophore (Seglen et al, 1990), or isolation membrane ).…”