Hydrophobic ligand binding properties of the human lipocalin apolipoprotein M.
Apolipoprotein M (apoM) is a plasma protein associated mainly with HDL. ApoM is suggested to be important for the formation of prebeta-HDL, but its mechanism of action is unknown. Homology modeling has suggested apoM to be a lipocalin. Lipocalins share a structurally conserved beta-barrel, which in many lipocalins bind hydrophobic ligands. The aim of this study was to test the ability of apoM to bind different hydrophobic substances. ApoM was produced both in Escherichia coli and in HEK 293 cells. Characterization of both variants with electrophoretic and immunological methods suggested apoM from E. coli to be correctly folded. Intrinsic tryptophan fluorescence of both apoM variants revealed that retinol, all-trans-retinoic acid, and 9-cis-retinoic acid bound (dissociation constant = 2-3 microM), whereas other tested substances (e.g., cholesterol, vitamin K, and arachidonic acid) did not. The intrinsic fluorescence of two apoM mutants carrying single tryptophans was quenched by retinol and retinoic acid to the same extent as wild-type apoM, indicating that the environment of both tryptophans was affected by the binding. In conclusion, the binding of retinol and retinoic acid supports the hypothesis that apoM is a lipocalin. The physiological relevance of this binding has yet to be elucidated.
Ahnström
Josefin
J
Department of Laboratory Medicine, Division of Clinical Chemistry, Lund University, University Hospital, SE-20502 Malmö, Sweden.
Faber
Kirsten
K
Axler
Olof
O
Dahlbäck
Björn
B
D000817:Anilino Naphthalenesulfonates / Q000737:chemistry / Q000378:metabolism; D000911:Antibodies, Monoclonal / Q000378:metabolism; D001053:Apolipoproteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D000075243:Apolipoproteins M; D001665:Binding Sites; D002352:Carrier Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D002478:Cells, Cultured; D004797:Enzyme-Linked Immunosorbent Assay; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D006801:Humans; D057927:Hydrophobic and Hydrophilic Interactions; D008024:Ligands; D054834:Lipocalins; D008958:Models, Molecular; D011487:Protein Conformation; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D013050:Spectrometry, Fluorescence; D014162:Transfection; D014212:Tretinoin / Q000737:chemistry / Q000378:metabolism*; D014801:Vitamin A / Q000737:chemistry / Q000378:metabolism*
C400523:APOM protein, human; D000817:Anilino Naphthalenesulfonates; D000911:Antibodies, Monoclonal; D001053:Apolipoproteins; D000075243:Apolipoproteins M; D002352:Carrier Proteins; D008024:Ligands; D054834:Lipocalins; D011994:Recombinant Proteins; D014801:Vitamin A; D014212:Tretinoin; C027132:1-anilino-8-naphthalenesulfonate