Photoaffinity labeling and fatty acid permeation in 3T3-L1 adipocytes

• Published in: The Journal of biological chemistry Vol. 266; no. 33; pp. 22621 - 22625
• Main Authors: TRIGATTI, B. L, MANGROO, D, GERBER, G. E
• Format: Journal Article
• Language: English
• Published: Bethesda, MD American Society for Biochemistry and Molecular Biology 25.11.1991
• Subjects: 3T3 Cells; Adipose Tissue - metabolism; Analytical, structural and metabolic biochemistry; Animals; Azirines - metabolism; Binding and carrier proteins; Biological and medical sciences; Biological Transport; Carrier Proteins - isolation & purification; Carrier Proteins - metabolism; Cell Differentiation; Cell Membrane - metabolism; cell membranes; Fatty Acid-Binding Protein 7; Fatty Acid-Binding Proteins; fatty acids; Fatty Acids, Nonesterified - metabolism; Fundamental and applied biological sciences. Psychology; Kinetics; Mice; Molecular Weight; Neoplasm Proteins; Nerve Tissue Proteins; Oleic Acid; Oleic Acids - metabolism; Proteins; Characterization; Adipocyte; Binding protein; Oleic acid; Plasma membrane; 3T3-Cell line; Photoaffinity labelling; Cell differentiation; Fatty acids
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1016/s0021-9258(18)54616-0

Long chain fatty acid uptake was investigated in 3T3-L1 cells. Differentiation of these cells from fibroblasts to adipocytes was accompanied by an 8.5-fold increase in the rate of oleate uptake. This was saturable in adipocytes with apparent Kt and Vmax values of 78 nM and 16 nmol/min/mg cell protein, respectively. A number of proteins in various subcellular fractions of differentiated cells were labeled with the photoreactive fatty acid 11-m-diazirinophenoxy[11-3H]undecanoate. A 15-kDa cytoplasmic protein was induced upon differentiation to adipocytes. This protein was labeled with the photoreactive fatty acid in cytoplasm isolated from differentiated adipocytes, but not in cytoplasm from undifferentiated, fibroblastic cells. Furthermore, a high affinity fatty acid binding protein of 22 kDa was identified in plasma membranes of undifferentiated cells, and its level of labeling increased 2-fold upon differentiation. These results indicate the usefulness of the photoreactive fatty acid in identifying cellular fatty acid binding proteins, and its potential to elucidate the spatial and temporal distribution of fatty acids in intact cells.