Peroxisomal membrane protein PMP68 of mouse liver: cloning of a cDNA encompassing the nucleotide binding fold and epitope mapping of monoclonal antibodies to the expressed protein.
We have isolated and sequenced a cDNA which encodes 376 amino acids toward the carboxy-terminus, and encompassing the putative nucleotide binding fold, of PMP68 (mouse liver peroxisomal integral membrane protein of 68 kDa) the major integral membrane protein of mouse liver peroxisomes. The protein sequence predicted from this cDNA shows 97.9% amino acid identity to this same region of rat liver PMP70, a member of the ATP-binding cassette protein superfamily (K. Kamijo, S. Taketani, S. Yokota, T. Osumi, and T. Hashimoto, 1990, J. Biol. Chem. 265, 4534-4540). The section of the cDNA encoding the hydrophilic and putative cytoplasmic domain of PMP68 was expressed as a recombinant fusion protein in bacteria. Two monoclonal antibodies raised against this protein have been epitope-mapped to peptides generated by cyanogen bromide cleavage of the fusion protein. Antibody 1A4 recognizes a peptide whose sequence contains the first motif of the putative nucleotide binding fold of PMP68, and antibody 8F11 recognizes a carboxy-terminal peptide which includes the second motif of this nucleotide binding fold. These antibodies are expected to be useful in the elucidation of the biological function of this putative membrane transporter.
Chen
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Faculty of Science and Technology, Griffith University, Nathan, Brisbane, Australia.
Lu
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Land
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Ayres
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Crane
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D018528:ATP-Binding Cassette Transporters; D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal; D001483:Base Sequence; D001665:Binding Sites; D002850:Chromatography, Gel; D002851:Chromatography, High Pressure Liquid; D003001:Cloning, Molecular; D018076:DNA, Complementary; D004591:Electrophoresis, Polyacrylamide Gel; D000939:Epitopes / Q000032:analysis*; D005260:Female; D015151:Immunoblotting; D008099:Liver / Q000378:metabolism*; D008565:Membrane Proteins / Q000096:biosynthesis* / Q000737:chemistry* / Q000378:metabolism; D051379:Mice; D008817:Mice, Mutant Strains; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000737:chemistry / Q000302:isolation & purification; D017510:Protein Folding; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000378:metabolism
D018528:ATP-Binding Cassette Transporters; C489271:Abcd3 protein, mouse; C489272:Abcd3 protein, rat; D000911:Antibodies, Monoclonal; D018076:DNA, Complementary; D000939:Epitopes; D008565:Membrane Proteins; D010446:Peptide Fragments; D011993:Recombinant Fusion Proteins