Purification and characterization of fatty acid-binding proteins from brown adipose tissue of the rat.
Fatty acid-binding proteins (FABPs) have been identified and purified from interscapular brown adipose tissue of the rat. The proteins were characterized and their properties compared with the FABP present in white adipose tissue. FABP was purified to electrophoretic homogeneity from brown adipose tissue by a procedure involving precipitation with 70% ammonium sulphate, followed sequentially by ion-exchange chromatography and gel filtration chromatography. The purified fraction migrated as a single band on SDS-PAGE with an apparent molecular mass of 14,200. Scatchard analysis of [14C]oleate-binding to purified FABP gave a Kd value of approx. 0.80 +/- 0.02 microM and a maximal binding of 0.65 +/- 0.03 mol per mol of protein; these values were similar to that found with the FABP purified from white fat. The FABP concentration in brown adipose tissue was almost twice that of FABP in white adipose tissue. Fatty acid analysis of FABP from brown adipose tissue revealed that the intrinsic arachidonic acid content was proportionately higher than that present in FABP of white adipose tissue. Isoelectric focusing of delipidated FABP indicated that it existed with two charge isoforms (pI 6.85 and 7.35). The purified FABP additionally emerged in two peaks (FABP-I and FABP-II) from a reverse phase HPLC column. Amino acid analysis showed that Gly, Thr, and Ser residues in FABP-I were almost twice as high as in FABP-II. The N-terminals of both FABP-I and -II were not blocked. These components have been partially sequenced and showed a sequence homology only between 25-31 residues from the N-terminal. Further studies are required to elucidate the precise function of the two different isoforms of FABP in brown adipose tissue.
Dutta-Roy
A K
AK
Division of Biochemical Sciences, Rowett Research Institute, Aberdeen, UK.
Huang
Y
Y
Dunbar
B
B
Trayhurn
P
P
D002001:Adipose Tissue, Brown / Q000737:chemistry*; D000595:Amino Acid Sequence; D000596:Amino Acids / Q000032:analysis; D000818:Animals; D016718:Arachidonic Acid / Q000032:analysis; D002352:Carrier Proteins / Q000737:chemistry / Q000302:isolation & purification*; D000071438:Fatty Acid-Binding Protein 7; D050556:Fatty Acid-Binding Proteins; D007525:Isoelectric Focusing; D019787:Linoleic Acid; D008041:Linoleic Acids / Q000032:analysis; D008297:Male; D008969:Molecular Sequence Data; D009363:Neoplasm Proteins; D009419:Nerve Tissue Proteins; D019308:Palmitic Acid; D010169:Palmitic Acids / Q000032:analysis; D051381:Rats
D000596:Amino Acids; D002352:Carrier Proteins; C490174:Fabp7 protein, rat; D000071438:Fatty Acid-Binding Protein 7; D050556:Fatty Acid-Binding Proteins; D008041:Linoleic Acids; D009363:Neoplasm Proteins; D009419:Nerve Tissue Proteins; D010169:Palmitic Acids; D016718:Arachidonic Acid; D019308:Palmitic Acid; D019787:Linoleic Acid