Purification and properties of Yip3/PRA1 as a Rab GDI displacement factor.
Prenylated Rab proteins exist in the cytosol bound to guanine dissociation inhibitor (GDI). These dimeric complexes contain all of the information needed for accurate membrane delivery. We have shown that membranes contain a proteinaceous activity that is required for Rab delivery, and we named that activity GDI displacement factor (GDF). Biochemical analysis revealed that GDF activity was membrane associated and had a mass of approximately 25 kDa. We therefore used a candidate gene approach and were able to show that pure Yip3/PRA1 protein displays GDF activity. In this chapter, we review key aspects of GDF analysis: our assay and the method by which we purify Yip3/PRA1 in active form.
Sivars
Ulf
U
Aivazian
Dikran
D
Pfeffer
Suzanne
S
D000595:Amino Acid Sequence; D002853:Chromatography, Liquid; D004591:Electrophoresis, Polyacrylamide Gel; D020730:Guanine Nucleotide Dissociation Inhibitors / Q000737:chemistry / Q000302:isolation & purification* / Q000502:physiology*; D008969:Molecular Sequence Data; D011994:Recombinant Proteins / Q000737:chemistry / Q000302:isolation & purification; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry / Q000302:isolation & purification* / Q000502:physiology*; D033921:Vesicular Transport Proteins / Q000737:chemistry / Q000302:isolation & purification* / Q000502:physiology*
C086796:GDP dissociation inhibitor 1; D020730:Guanine Nucleotide Dissociation Inhibitors; D011994:Recombinant Proteins; D029701:Saccharomyces cerevisiae Proteins; D033921:Vesicular Transport Proteins; C505724:Yip3 protein, S cerevisiae