Golgins protein

They are designated golgin-84, -95, -160, -245, and -376 (giantin or macrogolgin) and were identified initially as human autoantigens (Chapter 31), appearing in the blood of persons with autoimmune disorders such as Sjogren s syndrome.281 282 Another protein... 

Diao, A., Rahman, D., Pappin, D.J., Lucocq, J. and Lowe, M. (2003) The coiled-coil membrane protein golgin-84 is a novel rab effector required for Golgi ribbon formation. J. Cell Biol. 160, 201-212. 

Instead of biotinylation, fluorescently labeled COPI vesicles can be purified using this method. However, reagents such as NHS-Alexa-488 (Molecular Probes, A-10235) label many proteins in the COPI vesicles in addition to golgin-84. 

Bascom, R. A, Srinivasan, S., and Nussbaum, R. L. (1999). Identification and characterization of golgin-84, a novel Golgi integral membrane protein with a cytoplasmic coiled-coil domain. J. Biol Chem. 274, 2953-2962. 

Shorter, J., Beard, M. B., Seemann, J., Dirac-Svejstrup, A. B., and Warren, G. (2002). Sequential tethering of Golgins and catalysis of SNAREpin assembly by the vesicle-tethering protein pll5. J. Cell Biol 157, 45-62. 

In the pull-down experiments using recombinant protein, the GRIP domain of golgin-245 interacts selectively with GTP-bound Aril, thns supporting the proposal that golgin-245 is an effector of Aril in vivo. This interaction is direct as shown recently by our X-ray crystal structure of the GRIP domain complexed with Arll(GTP) (Wu et al., 2004). 

The in vitro transport assay is routinely characterized to ensure that the assay has indeed faithfully reconstituted the EE/RE-to-TGN transport in vitro (Tai et ah, 2004). First, the assay must be cytosol-dependent. The basal level of transport in the absence of cytosol is usually 5-10% of that in the presence of 1.2 mg/ml of rat liver cytosol. Second, the assay is tested with some commonly used inhibitors such as Brefeldin A and guanosine 5 -0-(3-thio) triphosphate (GTP7S), as their inhibitory role in EE/RE-to-TGN traffic is well documented. Finally, the assay is tested using antibodies (such as autibody agaiust syutaxiulb) or recombinant proteins (such as GST-GRIP of golgin-97) that have been shown to inhibit in vitro EE/RE-to-TGN transport of STxB. 

Fig. 1. The recombinant GRIP domain of golgin-97 inhibits in vitro EE/RE-TGN transport. STxB transport assays are carried out in the absence (lanes 1 and 4) or presence of 133 g/ml of GST-GRIP domain proteins (lane 2, GST-GRIP lane 3, heat-denatured GST-GRIP lane 5, GST-GRIPA 697A). Extent of transport relative to controls (set as 100%) is calculated from two to three independent experiments. Error bars represent standard deviations. Bottom panel shows representative phosphor-imaging data. Reprinted from Molecular Biology of the Cell (Mol. Biol. Cell (2004). 15, 4426-4443.) with the permission of The American Society for Cell Biology.

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