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J. Biol. Chem., Vol. 275, Issue 33, 25672-25680, August 18, 2000
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From the Vitamin E (
A Novel Human Tocopherol-associated Protein
CLONING, IN VITRO EXPRESSION, AND
CHARACTERIZATION*
§,§¶,
,,
Institute of Biochemistry and Molecular
Biology, University of Bern, Bühlstrasse 28, 3012 Bern,
Switzerland, the Institute of Biochemistry & Biophysics, Tehran
University, P. O. Box 13145-1384, Tehran, Iran, and the ** Institute for
General Microbiology, University of Bern, Balzerstrasse 4, 3012 Bern,
Switzerland
-tocopherol) is an essential
dietary nutrient for humans and animals. The mechanisms involved in
cellular regulation as well as in the preferential cellular and tissue
accumulation of -tocopherol are not yet well established. We
previously reported (Stocker, A., Zimmer, S., Spycher, S. E., and
Azzi, A. (1999) IUBMB Life 48, 49-55) the identification
of a novel 46-kDa tocopherol-associated protein (TAP) in the cytosol of
bovine liver. Here, we describe the identification, the molecular
cloning into Escherichia coli, and the in vitro
expression of the human homologue of bovine TAP, hTAP. This protein appears to belong to a family of
hydrophobic ligand binding proteins, which have the CRAL
(cis-retinal binding motif) sequence in common. By using a
biotinylated -tocopherol derivative and the IASys resonant mirror
biosensor, the purified recombinant protein was shown to bind
tocopherol at a specific binding site with Kd
4.6 × 10
7 M. Northern analyses showed
that hTAP mRNA has a size of approximately 2800 base pairs and is
ubiquitously expressed. The highest amounts of hTAP message are found
in liver, brain, and prostate. In conclusion, hTAP has sequence
homology to proteins containing the CRAL_TRIO structural
motif. TAP binds to -tocopherol and biotinylated tocopherol, suggesting the existence of a hydrophobic pocket, possibly analogous to
that of SEC14.
*
This study was made possible by the support of the Swiss
National Science Foundation, by a grant of the Henkel Corporation, and
by the Stiftung für die Ernährungsforschung in der Schweiz.The costs of publication of this
article were defrayed in part by the
payment of page charges. The article
must therefore be hereby marked
"advertisement" in
accordance with 18 U.S.C. Section
1734 solely to indicate this fact.
Copyright © 2000 by The American Society for Biochemistry and Molecular Biology, Inc.
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