Membrane topology of the ATP binding cassette transporter ABCR and its relationship to ABC1 and related ABCA transporters: identification of N-linked glycosylation sites.
ABCR is a member of the ABCA subclass of ATP binding cassette transporters that is responsible for Stargardt macular disease and implicated in retinal transport across photoreceptor disc membranes. It consists of a single polypeptide chain arranged in two tandem halves, each having a multi-spanning membrane domain followed by a nucleotide binding domain. To delineate between several proposed membrane topological models, we have identified the exocytoplasmic (extracellular/lumen) N-linked glycosylation sites on ABCR. Using trypsin digestion, site-directed mutagenesis, concanavalin A binding, and endoglycosidase digestion, we show that ABCR contains eight glycosylation sites. Four sites reside in a 600-amino acid exocytoplasmic domain of the N-terminal half between the first transmembrane segment H1 and the first multi-spanning membrane domain, and four sites are in a 275-amino acid domain of the C half between transmembrane segment H7 and the second multi-spanning membrane domain. This leads to a model in which each half has a transmembrane segment followed by a large exocytoplasmic domain, a multi-spanning membrane domain, and a nucleotide binding domain. Other ABCA transporters, including ABC1 linked to Tangier disease, are proposed to have a similar membrane topology based on sequence similarity to ABCR. Studies also suggest that the N and C halves of ABCR are linked through disulfide bonds.
Bungert
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Departments of Biochemistry & Molecular Biology and Ophthalmology, University of British Columbia, Vancouver, British Columbia V6T 1Z3, Canada.
Molday
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Molday
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D064286:ATP Binding Cassette Transporter 1; D018528:ATP-Binding Cassette Transporters / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D000595:Amino Acid Sequence; D000818:Animals; D019556:COS Cells; D002417:Cattle; D003208:Concanavalin A / Q000378:metabolism; D016384:Consensus Sequence; D004220:Disulfides / Q000378:metabolism; D006023:Glycoproteins / Q000737:chemistry*; D006031:Glycosylation; D006801:Humans; D008954:Models, Biological; D008969:Molecular Sequence Data; D017434:Protein Structure, Tertiary; D012374:Rod Cell Outer Segment / Q000378:metabolism; D017384:Sequence Deletion; D017386:Sequence Homology, Amino Acid; D014162:Transfection; D014357:Trypsin / Q000737:chemistry
C577836:ABCA1 protein, human; C056475:ABCA4 protein, human; D064286:ATP Binding Cassette Transporter 1; D018528:ATP-Binding Cassette Transporters; D004220:Disulfides; D006023:Glycoproteins; D003208:Concanavalin A; D014357:Trypsin