title: Arabidopsis PEX19 is a dimeric protein that binds the peroxin PEX10.
abstract: Peroxisomes are organelles found in all eukaryotic cells. Peroxisomes import integral membrane proteins post-translationally, and PEX19 is a predominantly cytosolic, farnesylated protein of mammalian and yeast cells that binds multiple peroxisome membrane proteins and is required for their correct targeting/insertion to the peroxisome membrane. We report the characterisation of the Arabidopsisthaliana homologue of PEX19 which is a predominantly cytosolic protein. AtPEX19 is encoded by two genes (designated AtPEX19-1 and AtPEX19-2) that are expressed in all tissues and at all developmental stages of the plant. Quantitative real time PCR shows that AtPEX19-1 and AtPEX19-2 have distinct expression profiles. Using in vitro translation and co-immunoprecipitation AtPEX19-1 was shown to bind to the Arabidopsis peroxisomal membrane protein PEX10. Additionally, bacterially expressed recombinant AtPEX19-1 was able to bind a fusion protein consisting of the C-terminus of PEX10 and glutathione S-transferase in pull-down assays, thereby demonstrating that non-farnesylated AtPEX19 can interact with the C-terminus of AtPEX10. Purified recombinant AtPEX19-1 was analysed by gel filtration chromatography and was found to have a molecular weight consistent with it forming a dimer and a dimer was detected in Arabidopsis cell extracts that was slightly destabilised in the presence of DTT. Moreover, cross-linking studies of native AtPEX19 suggest that in vivo it is the dimeric species of the protein that preferentially forms complexes with other proteins.
authors.affiliation: Biosciences, Sheffield Hallam University, Sheffield, S1 1WB, UK. d.hadden@shu.ac.uk
authors.forename: Dawn A
authors.initials: DA
authors.lastname: Hadden
authors.forename: Belinda A
authors.initials: BA
authors.lastname: Phillipson
authors.forename: Katherine A
authors.initials: KA
authors.lastname: Johnston
authors.forename: Laura-Anne
authors.initials: LA
authors.lastname: Brown
authors.forename: Iain W
authors.initials: IW
authors.lastname: Manfield
authors.forename: Mahmoud
authors.initials: M
authors.lastname: El-Shami
authors.forename: Imogen A
authors.initials: IA
authors.lastname: Sparkes
authors.forename: Alison
authors.initials: A
authors.lastname: Baker
mesh_terms: D017360:Arabidopsis / Q000378:metabolism*; D029681:Arabidopsis Proteins / Q000235:genetics / Q000378:metabolism*; D003600:Cytosol / Q000378:metabolism; D019281:Dimerization; D017343:Genes, Plant; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism*; D026901:Membrane Transport Proteins / Q000378:metabolism*; D000074401:Peroxins; D020675:Peroxisomes / Q000378:metabolism
chemical_list: D029681:Arabidopsis Proteins; D008565:Membrane Proteins; D026901:Membrane Transport Proteins; C477605:PEX10 protein, Arabidopsis; C514419:PEX19 protein, Arabidopsis; D000074401:Peroxins