A glycoprotein binding retinoids and fatty acids is present in Drosophila.
In the search for a possible Drosophila melanogaster homolog of interphotoreceptor retinoid-binding protein (IRBP), a approximately 140-kDa retinoid- and fatty acid-binding glycoprotein found in vertebrates, the 110,000 g supernatant fraction prepared from homogenates of fly heads was analyzed for the presence of proteins capable of binding radiolabeled retinol and palmitic acid. A soluble protein, which binds concanavalin A and has a retention time on size-exclusion high-performance liquid chromatography identical to that of purified bovine IRBP, was identified as binding both ligands. As assessed by fluorescence titration, the protein fraction obtained by concanavalin A-Sepharose affinity chromatography and size-exclusion chromatography of fly head supernatant had apparent dissociation constants of 2.9 x 10(-7) +/- 0.6 M for all-trans retinol, with the number (n) of independent ligand binding sites per protein molecule = 2, and 3.5 x 10(-7) +/- 0.1 M for 16-[9-anthroyloxy] palmitic acid with n = 7. High-performance liquid chromatography of hexane extracts of this protein fraction resolved several peaks with polarity and relative retention times similar, but not identical to all-trans retinol and retinal and their 9-, 11-, and 13-cis isomers. Gas chromatography/mass spectrometry analysis of fatty acid methyl esters prepared following lipid extraction of the protein identified lauric, myristic, palmitic, palmitoleic, and oleic acids as being covalently bound. Laurate, myristate, palmitate, and stearate were noncovalently bound. The apparent molecular mass of the Drosophila protein as assessed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and silver staining of the retinoid- and fatty acid-binding peak obtained by hydrophobic interaction chromatography of the size-exclusion fraction was approximately 70 kDa.
Duncan
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T
Laboratory of Retinal Cell and Molecular Biology, National Eye Institute, National Institutes of Health, Bethesda, Maryland 20892.
Kutty
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G
Chader
G J
GJ
Wiggert
B
B
D000818:Animals; D001665:Binding Sites; D002851:Chromatography, High Pressure Liquid; D003208:Concanavalin A / Q000378:metabolism; D004331:Drosophila melanogaster / Q000737:chemistry*; D005136:Eye Proteins; D005229:Fatty Acids, Monounsaturated / Q000032:analysis; D006023:Glycoproteins / Q000378:metabolism*; D006257:Head; D007850:Lauric Acids / Q000032:analysis; D008024:Ligands; D019814:Myristic Acid; D009227:Myristic Acids / Q000032:analysis; D019308:Palmitic Acid; D010169:Palmitic Acids / Q000032:analysis / Q000378:metabolism*; D012177:Retinol-Binding Proteins / Q000737:chemistry*; D014801:Vitamin A / Q000378:metabolism*; D000099214:Retinol-Binding Proteins, Interstitial
D003208:Concanavalin A; D005136:Eye Proteins; D005229:Fatty Acids, Monounsaturated; D006023:Glycoproteins; D007850:Lauric Acids; D008024:Ligands; D019814:Myristic Acid; D009227:Myristic Acids; D019308:Palmitic Acid; D010169:Palmitic Acids; D012177:Retinol-Binding Proteins; D014801:Vitamin A; D000099214:Retinol-Binding Proteins, Interstitial; C030358:lauric acid; C008757:palmitoleic acid; C047265:16-(9-anthroyloxy)palmitic acid