High-affinity fatty acid-binding activity in epidermis and cultured keratinocytes is attributable to high-molecular-weight and not low-molecular-weight fatty acid-binding proteins.
Fatty acid-binding proteins (FABPs) are abundant low-molecular-weight cytosolic proteins in tissues involved in fatty acid (FA) metabolism. Because epidermis is also an active lipogenic tissue, we examined cytosols from murine and porcine epidermis and cultured human keratinocytes and fibroblasts for FABPs. High-affinity FA-binding activity was present in both epidermis and differentiated keratinocytes, whereas no high-affinity FA-binding activity was found in cultured human fibroblasts or undifferentiated keratinocytes. By column chromatography, a single binding peak was identified in the high (90-100 kDa)-molecular-weight range and no binding activity was evident in the low (14-15 kDa)-molecular-weight range, where conventional FABPs elute. Moreover, rabbit anti-rat heart FABP, anti-rat intestine FABP, and anti-rat liver FABP antisera did not identify proteins in the 14-15-kDa range in murine epidermal cytosol by Western immunoblots, whereas the anti-rat-heart antibody recognized a protein of approximately 32 kDa. Isoelectric focusing of differentiated keratinocyte cytosol demonstrated a single FA-binding peak having a pI of approximately 4.0. Analysis of this binding peak by SDS-PAGE revealed peptides of approximately 66 and 38 kDa. These findings suggest the possibility that the FA-binding protein in keratinocyte cytosol normally exists as a heterodimer. Western immunoblots of both differentiated keratinocyte cytosol and keratinocyte-conditional media stained with a rabbit anti-human serum albumin antibody identified a protein of approximately 67 kDa, but the electrofocused fraction did not react with this antibody. Thus, epidermis and differentiated keratinocytes possess high-affinity cytosolic FA-binding activity that cannot be ascribed either to conventional low-molecular-weight FABPs or to albumin.
Schurer
N Y
NY
Dermatology Services, Department of Veterans Affairs Medical Center, San Francisco, California 94121.
Bass
N M
NM
Jin
S
S
Manning
J A
JA
Pillai
S
S
Williams
M L
ML
D001667:Binding, Competitive; D015153:Blotting, Western; D002352:Carrier Proteins / Q000737:chemistry / Q000378:metabolism*; D002478:Cells, Cultured; D002850:Chromatography, Gel; D003600:Cytosol / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D004817:Epidermis / Q000378:metabolism*; D000071438:Fatty Acid-Binding Protein 7; D050556:Fatty Acid-Binding Proteins; D005227:Fatty Acids / Q000378:metabolism*; D006801:Humans; D007525:Isoelectric Focusing; D015603:Keratinocytes / Q000378:metabolism*; D008970:Molecular Weight; D009363:Neoplasm Proteins; D009419:Nerve Tissue Proteins; D025521:Tumor Suppressor Proteins
D002352:Carrier Proteins; C107608:FABP7 protein, human; C490174:Fabp7 protein, rat; D000071438:Fatty Acid-Binding Protein 7; D050556:Fatty Acid-Binding Proteins; D005227:Fatty Acids; D009363:Neoplasm Proteins; D009419:Nerve Tissue Proteins; D025521:Tumor Suppressor Proteins