Reversible dissociation of coatomer: functional characterization of a beta/delta-coat protein subcomplex.
COPI-coated vesicles mediate protein transport within the early secretory pathway. Their coat consists of ADP ribosylation factor (ARF1, a small guanosine nucleotide binding protein), and coatomer, a cytosolic complex composed of seven subunits, alpha- to zeta-coat proteins (COPs). For coat formation that initiates budding of a vesicle, ARF1 is recruited to the Golgi membrane from the cytosol in its GTP-bound form, and subsequently, coatomer can bind to the membrane. To identify a minimal structure of coatomer capable to bind to Golgi membranes in an ARF1-dependent manner, we have established a procedure to dissociate coatomer under conditions that allow reassociation of the subunits to a complete and functional complex. After dissociation, subunits or subcomplexes can be isolated and may be expected to be functional. Herein we describe isolation of a subcomplex of coatomer consisting of beta- and delta-COPs that is able to bind to Golgi membranes in an ARF1- and GTP-dependent manner.
Pavel
J
J
Biochemie-Zentrum Heidelberg Ruprecht-Karls-Universität, Im Neuenheimer Feld 328, 69120 Heidelberg, Germany.
Harter
C
C
Wieland
F T
FT
D020823:ADP-Ribosylation Factor 1; D020727:ADP-Ribosylation Factors; D000818:Animals; D020756:Coatomer Protein; D003600:Cytosol / Q000378:metabolism; D019281:Dimerization; D019204:GTP-Binding Proteins / Q000378:metabolism*; D006056:Golgi Apparatus / Q000378:metabolism; D016244:Guanosine 5'-O-(3-Thiotriphosphate) / Q000494:pharmacology; D006153:Guanosine Diphosphate / Q000031:analogs & derivatives / Q000494:pharmacology; D006160:Guanosine Triphosphate / Q000378:metabolism; D008099:Liver / Q000378:metabolism; D046911:Macromolecular Substances; D008299:Maleic Anhydrides / Q000494:pharmacology; D008565:Membrane Proteins / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism*; D008869:Microtubule-Associated Proteins / Q000302:isolation & purification / Q000378:metabolism*; D011817:Rabbits; D013873:Thionucleotides / Q000494:pharmacology
D020756:Coatomer Protein; D046911:Macromolecular Substances; D008299:Maleic Anhydrides; D008565:Membrane Proteins; D008869:Microtubule-Associated Proteins; D013873:Thionucleotides; D006153:Guanosine Diphosphate; D016244:Guanosine 5'-O-(3-Thiotriphosphate); C007474:2,3-dimethylmaleic anhydride; C023427:guanosine 5'-O-(2-thiodiphosphate); D006160:Guanosine Triphosphate; D019204:GTP-Binding Proteins; D020823:ADP-Ribosylation Factor 1; D020727:ADP-Ribosylation Factors