Characterisation of the Rab binding properties of Rab coupling protein (RCP) by site-directed mutagenesis.
Rab coupling protein (RCP) is a member of the Rab11-family of interacting proteins (Rab11-FIPs). Family members are characterised by their ability to interact with Rab11. This property is mediated by a conserved Rab binding domain (RBD) located at their carboxy-termini. Several Rab11-FIPs can also interact with other small GTPases. RCP interacts with Rab4 in addition to Rab11. To dissect out the individual properties of the Rab4 and Rab11 interactions with RCP, conserved amino acids within the RBD of RCP were mutated by site-directed mutagenesis. The effect of these mutations on Rab4 and Rab11 binding, and the intracellular localisation of RCP, was examined. Our results indicate that Rab11, rather than Rab4, mediates the intracellular localisation of RCP, and that the class I Rab11-FIPs compete for binding to Rab11.
Molecular Cell Biology Laboratory, Department of Biochemistry, Biosciences Institute, University College Cork, Cork, Ireland.
Andrew J
AJ
Lindsay
Mary W
MW
McCaffrey
D048868:Adaptor Proteins, Signal Transducing; D019883:Alkyl and Aryl Transferases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D000595:Amino Acid Sequence; D001483:Base Sequence; D001665:Binding Sites; D002352:Carrier Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D045744:Cell Line, Tumor; D017931:DNA Primers; D006367:HeLa Cells; D006801:Humans; D008565:Membrane Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D010957:Plasmids; D011485:Protein Binding; D011487:Protein Conformation; D011994:Recombinant Proteins / Q000378:metabolism; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D020691:rab GTP-Binding Proteins / Q000378:metabolism*
D048868:Adaptor Proteins, Signal Transducing; C081787:CHM protein, human; D002352:Carrier Proteins; D017931:DNA Primers; D008565:Membrane Proteins; C488966:RAB11FIP1 protein, human; D011994:Recombinant Proteins; D019883:Alkyl and Aryl Transferases; D020691:rab GTP-Binding Proteins