title: Structural requirements for ligand binding by a probable plant vacuolar sorting receptor.
abstract: How sorting receptors recognize amino acid determinants on polypeptide ligands and respond to pH changes for ligand binding or release is unknown. The plant vacuolar sorting receptor BP-80 binds polypeptide ligands with a central Asn-Pro-Ile-Arg (NPIR) motif. tBP-80, a soluble form of the receptor lacking transmembrane and cytoplasmic sequences, binds the peptide SSSFADSNPIRPVTDRAASTYC as a monomer with a specificity indistinguishable from that of BP-80. tBP-80 contains an N-terminal region homologous to ReMembR-H2 (RMR) protein lumenal domains, a unique central region, and three C-terminal epidermal growth factor (EGF) repeats. By protease digestion of purified secreted tBP-80, and from ligand binding studies with a secreted protein lacking the EGF repeats, we defined three protease-resistant structural domains: an N-terminal/RMR homology domain connected to a central domain, which together determine the NPIR-specific ligand binding site, and a C-terminal EGF repeat domain that alters the conformation of the other two domains to enhance ligand binding. A fragment representing the central domain plus the C-terminal domain could bind ligand but was not specific for NPIR. These results indicate that two tBP-80 binding sites recognize two separate ligand determinants: a non-NPIR site defined by the central domain-EGF repeat domain structure and an NPIR-specific site contributed by the interaction of the N-terminal/RMR homology domain and the central domain.
authors.affiliation: Institute of Biological Chemistry, Washington State University, Pullman, Washington 99164-6340, USA.
authors.forename: X
authors.initials: X
authors.lastname: Cao
authors.forename: S W
authors.initials: SW
authors.lastname: Rogers
authors.forename: J
authors.initials: J
authors.lastname: Butler
authors.forename: L
authors.initials: L
authors.lastname: Beevers
authors.forename: J C
authors.initials: JC
authors.lastname: Rogers
mesh_terms: D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology / Q000494:pharmacology; D001665:Binding Sites; D002460:Cell Line; D003546:Cysteine Endopeptidases / Q000737:chemistry / Q000378:metabolism; D004220:Disulfides / Q000378:metabolism; D004331:Drosophila melanogaster; D010450:Endopeptidases / Q000378:metabolism; D004792:Enzyme Precursors / Q000737:chemistry / Q000378:metabolism; D004815:Epidermal Growth Factor / Q000737:chemistry; D005456:Fluorescent Dyes / Q000378:metabolism; D008024:Ligands; D008969:Molecular Sequence Data; D008970:Molecular Weight; D018532:Pisum sativum / Q000737:chemistry* / Q000235:genetics; D010446:Peptide Fragments / Q000737:chemistry / Q000378:metabolism; D010940:Plant Proteins; D011485:Protein Binding / Q000187:drug effects; D017434:Protein Structure, Tertiary; D011956:Receptors, Cell Surface / Q000737:chemistry* / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D020449:Repetitive Sequences, Amino Acid; D017384:Sequence Deletion / Q000235:genetics; D017386:Sequence Homology, Amino Acid; D013379:Substrate Specificity; D033921:Vesicular Transport Proteins
chemical_list: D000911:Antibodies, Monoclonal; D004220:Disulfides; D004792:Enzyme Precursors; D005456:Fluorescent Dyes; D008024:Ligands; D010446:Peptide Fragments; D010940:Plant Proteins; D011956:Receptors, Cell Surface; D033921:Vesicular Transport Proteins; D004815:Epidermal Growth Factor; D010450:Endopeptidases; D003546:Cysteine Endopeptidases; C076060:proaleurain