Purification of murine adipocyte lipid-binding protein. Characterization as a fatty acid- and retinoic acid-binding protein.
An adipose-specific protein has been purified from murine 3T3-L1 adipocytes to greater than 98% homogeneity. A purification procedure was developed utilizing a combination of gel filtration, cation exchange chromatography, and covalent chromatography on activated-thiol Sepharose 4B. The protein exists as a single polypeptide with a molecular weight of about 15,000, as determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The protein contains 2 mol of reduced sulfhydryl groups per mol of protein and an amino terminus blocked to sequencing. Automated Edman degradation of trypsin and CNBr-derived peptides has verified that the purified protein is that predicted by the mRNA (Bernlohr, D. A., Angus, C. W., Lane, M. D., Bolanowski, M. A., and Kelly, T. J. Jr. (1984) Proc. Natl. Acad. Sci. U.S.A. 81, 5468-5472). Based on sequence analysis, the 15-kDa adipocyte protein is considered to be a member of a family of tissue-specific, cytosolic lipid-binding proteins. Utilizing a liposome assay, the purified protein binds both oleic acid and retinoic acid saturably with approximately 1 mol of ligand bound per mol of protein. Dissociation constants determined from Scatchard analysis were 3 and 50 microM, respectively. This report represents the first demonstration of a member of this family of structurally related proteins that is capable of binding both fatty acid and retinoic acid. Hence, we propose the name adipocyte lipid-binding protein, or ALBP.
Matarese
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Department of Biochemistry, University of Minnesota, St. Paul 55108.
Bernlohr
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D000273:Adipose Tissue / Q000378:metabolism*; D000595:Amino Acid Sequence; D000818:Animals; D002352:Carrier Proteins / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism; D002478:Cells, Cultured; D002851:Chromatography, High Pressure Liquid; D000071438:Fatty Acid-Binding Protein 7; D050556:Fatty Acid-Binding Proteins; D005796:Genes; D007700:Kinetics; D051379:Mice; D008969:Molecular Sequence Data; D008970:Molecular Weight; D009363:Neoplasm Proteins; D009419:Nerve Tissue Proteins; D019301:Oleic Acid; D009829:Oleic Acids / Q000378:metabolism; D018168:Receptors, Retinoic Acid; D014212:Tretinoin / Q000378:metabolism
D002352:Carrier Proteins; C082667:Fabp5 protein, mouse; C086936:Fabp7 protein, mouse; D000071438:Fatty Acid-Binding Protein 7; D050556:Fatty Acid-Binding Proteins; D009363:Neoplasm Proteins; D009419:Nerve Tissue Proteins; D009829:Oleic Acids; D018168:Receptors, Retinoic Acid; D019301:Oleic Acid; D014212:Tretinoin