出版社：American Society for Biochemistry and Molecular Biology
摘要：Apolipoprotein M (apoM) is a plasma protein associated mainlywith HDL. ApoM is suggested to be important for the formationof preß-HDL, but its mechanism of action is unknown.Homology modeling has suggested apoM to be a lipocalin. Lipocalinsshare a structurally conserved ß-barrel, which inmany lipocalins bind hydrophobic ligands. The aim of this studywas to test the ability of apoM to bind different hydrophobicsubstances. ApoM was produced both in Escherichia coli and inHEK 293 cells. Characterization of both variants with electrophoreticand immunological methods suggested apoM from E. coli to becorrectly folded. Intrinsic tryptophan fluorescence of bothapoM variants revealed that retinol, all-trans-retinoic acid,and 9-cis-retinoic acid bound (dissociation constant = 2–3µM), whereas other tested substances (e.g., cholesterol,vitamin K, and arachidonic acid) did not. The intrinsic fluorescenceof two apoM mutants carrying single tryptophans was quenchedby retinol and retinoic acid to the same extent as wild-typeapoM, indicating that the environment of both tryptophans wasaffected by the binding. In conclusion, the binding of retinoland retinoic acid supports the hypothesis that apoM is a lipocalin.The physiological relevance of this binding has yet to be elucidated.Supplementary key words retinol • retinoic acid • intrinsic fluorescence • high density lipoprotein • signal peptide • cholesterol
Abbreviations: 1,8-ANS, 1-anilinonaphthalene-8-sulfonic acid; apoM, apolipoprotein M; Kd, dissociation constant; RBP, retinol binding protein
