title: Protein structure encodes the ligand binding specificity in pheromone binding proteins.
abstract: The ligand specificities and binding affinities of three recombinant pheromone binding proteins (PBPs) of two saturniid moths (genus Antheraea) were determined by using a novel binding assay in conjunction with two tritium-labeled constituents of the pheromone blend, [3H]-6E,11Z-hexadecadienyl acetate and [3H]-4E,9Z-tetradecadienyl acetate. The new binding assay, in which nonspecific adsorption to a plastic vessel is suppressed by presaturation of the surface with a 1-alkanol, allows measurement of dissociation constants (KD) for lipophilic ligands for their carrier proteins. The three PBPs showed KD values for [3H]-6E,11Z-16:Ac and [3H]-4E,9Z-14:Ac between 0.6 and 30 microM, as determined by Scatchard analysis. Importantly, two PBPs (Aper-1 and Aper-2) from one species showed opposite binding specificities for these two ligands. Aper-1, like Apol-3, showed 15-fold higher affinity for 6E,11Z-16:Ac than for 4E,9Z-14:Ac, while Aper-2 showed a 3.5-fold preference for binding the shorter chain compound. In addition, for the Apol-3 PBP, displacement of [3H]-6E,11Z-16:Ac binding by other pheromone components or analogs showed a clear trend in relative binding affinity: 6E,11Z-16:Ac > 4E,9Z-14:Ac > 6E,11Z-16:Al approximately 16:Ac > 6E,11Z-16:OH > 4E,9Z-14:OH. These data clearly demonstrate a > 1000-fold range of binding affinities among these very similar structures and unambiguously demonstrate the specificity of the PBP-pheromone interaction. Moreover, this assay offers the potential for determining ligand specificities for odorant binding proteins and other proteins in the vertebrate lipocalin superfamily.
authors.affiliation: Department of Chemistry, University at Stony Brook, New York 11794-3400, USA.
authors.forename: G
authors.initials: G
authors.lastname: Du
authors.forename: G D
authors.initials: GD
authors.lastname: Prestwich
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002352:Carrier Proteins / Q000737:chemistry / Q000378:metabolism*; D007301:Insect Hormones / Q000737:chemistry / Q000378:metabolism*; D019476:Insect Proteins; D036341:Intercellular Signaling Peptides and Proteins; D008969:Molecular Sequence Data; D009036:Moths; D009838:Oligodeoxyribonucleotides; D011487:Protein Conformation; D017386:Sequence Homology, Amino Acid
chemical_list: C094731:Aper-2 protein, Antheraea pernyi; D002352:Carrier Proteins; D007301:Insect Hormones; D019476:Insect Proteins; D036341:Intercellular Signaling Peptides and Proteins; D009838:Oligodeoxyribonucleotides; C067715:APR-1 pheromone-binding protein, Antheraea pernyi