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Hydrophobic Ligand Binding by Zn-α_2-glycoprotein, a Soluble Fat-depleting Factor Related to Major Histocompatibility Complex Proteins

Kennedy, Malcolm W. and Heikema, Astrid P. and Cooper, Alan and Bjorkman, Pamela J. and Sánchez, Luis M. (2001) Hydrophobic Ligand Binding by Zn-α_2-glycoprotein, a Soluble Fat-depleting Factor Related to Major Histocompatibility Complex Proteins. Journal of Biological Chemistry, 276 (37). pp. 35008-35013. ISSN 0021-9258. https://resolver.caltech.edu/CaltechAUTHORS:KENjbc01

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Abstract

Zn-alpha2-glycoprotein (ZAG) is a member of the major histocompatibility complex (MHC) class I family of proteins and is identical in amino acid sequence to a tumor-derived lipid-mobilizing factor associated with cachexia in cancer patients. ZAG is present in plasma and other body fluids, and its natural function, like leptin's, probably lies in lipid store homeostasis. X-ray crystallography has revealed an open groove between the helices of ZAG's alpha1 and alpha2 domains, containing an unidentified small ligand in a position similar to that of peptides in MHC proteins (Sanchez, L. M., Chirino, A. J., and Bjorkman, P. J. (1999) Science 283, 1914-1919). Here we show, using serum-derived and bacterial recombinant protein, that ZAG binds the fluorophore-tagged fatty acid 11-(dansylamino)undecanoic acid (DAUDA) and, by competition, natural fatty acids such as arachidonic, linolenic, eicosapentaenoic, and docosahexaenoic acids. Other MHC class I-related proteins (FcRn, HFE, HLA-Cw*0702) showed no such evidence of binding. Fluorescence and isothermal calorimetry analysis showed that ZAG binds DAUDA with Kd in the micromolar range, and differential scanning calorimetry showed that ligand binding increases the thermal stability of the protein. Addition of fatty acids to ZAG alters its intrinsic (tryptophan) fluorescence emission spectrum, providing a strong indication that ligand binds in the expected position close to a cluster of exposed tryptophan side chains in the groove. This study therefore shows that ZAG binds small hydrophobic ligands, that the natural ligand may be a polyunsaturated fatty acid, and provides a fluorescence-based method for investigating ZAG-ligand interactions.


Item Type:Article
ORCID:
AuthorORCID
Bjorkman, Pamela J.0000-0002-2277-3990
Alternate Title:Hydrophobic Ligand Binding by Zn-α2-glycoprotein, a Soluble Fat-depleting Factor Related to Major Histocompatibility Complex Proteins
Additional Information:© 2001 by The American Society for Biochemistry and Molecular Biology, Inc. Received for publication, June 6, 2001, and in revised form, June 24, 2001. Originally published In Press as doi:10.1074/jbc.C100301200 on June 25, 2001 We are indebted to Margaret Nutley and Fiona McMonagle for excellent technical help with the calorimetry experiments and protein preparation. This work was supported by The Wellcome Trust (United Kingdom) Grant 044156 (to M. W. K. and A. C.) and by the United States Army Medical Research and Material Command Breast Cancer Program. The biological microcalorimetry facilities are supported by the Biotechnology and Biological Sciences Research Council and Engineering and Physical Sciences Research Council (United Kingdom).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.
Funders:
Funding AgencyGrant Number
Wellcome Trust044156
Army Medical Research and Material Command Breast Cancer ProgramUNSPECIFIED
Biotechnology and Biological Sciences Research Council (BBSRC)UNSPECIFIED
Physical Sciences Research CouncilUNSPECIFIED
Issue or Number:37
Record Number:CaltechAUTHORS:KENjbc01
Persistent URL:https://resolver.caltech.edu/CaltechAUTHORS:KENjbc01
Alternative URL:http://dx.doi.org/10.1074/jbc.C100301200
Usage Policy:No commercial reproduction, distribution, display or performance rights in this work are provided.
ID Code:3921
Collection:CaltechAUTHORS
Deposited By: Archive Administrator
Deposited On:19 Jul 2006
Last Modified:30 Oct 2019 19:08

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