Localization of type I iodothyronine 5'-deiodinase to the basolateral plasma membrane in renal cortical epithelial cells
Type I iodothyronine 5'-deiodinase is an integral membrane protein catalyzing the phenolic ring deiodination of thyroxine. We recently showed that the substrate binding subunit of this approximately 50-kDa protein is selectively labeled with N-bromoacetyl-L-thyroxine, allowing ready identificat...
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Published in | The Journal of biological chemistry Vol. 266; no. 17; pp. 11262 - 11269 |
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Main Authors | , , , , |
Format | Journal Article |
Language | English |
Published |
Bethesda, MD
American Society for Biochemistry and Molecular Biology
15.06.1991
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Subjects | |
Online Access | Get full text |
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Summary: | Type I iodothyronine 5'-deiodinase is an integral membrane protein catalyzing the phenolic ring deiodination of thyroxine.
We recently showed that the substrate binding subunit of this approximately 50-kDa protein is selectively labeled with N-bromoacetyl-L-thyroxine,
allowing ready identification of the type I enzyme without the need to maintain catalytic activity. In this study, we used
both affinity labeling and catalytic activity to determine the regional distribution of this enzyme in rat kidney and to localize
the enzyme to specific plasma membrane domain(s) of renal epithelial cells. The type I enzyme was present exclusively in tubular
epithelial cells of the outer renal cortex and co-purified with basolateral plasma membranes; the renal medulla lacked activity.
LLC-PK1 cells, derived from the proximal convoluted tubule, have abundant type I 5'-deiodinating activity. We used this homogenous
cell line to verify that the type I enzyme was localized to the cytosolic surface of the basolateral membrane. Digitonin permeabilization
increased affinity labeling of the enzyme 4-fold, and approximately 75% of the affinity label was incorporated into the 27-kDa
substrate binding subunit. Affinity labeling of the type I enzyme in LLC-PK1 cells mimicked the affinity labeling of the substrate
binding subunit of type I 5'-deiodinase in rat kidney (Köhrle, J., Rasmussen, U. B., Ekenbarger, D. M., Alex, S., Rokos, H.,
Hesch, R. D., and Leonard, J. L. (1990) J. Biol. Chem. 265, 6155-6163). Subcellular fractionation of LLC-PK1 cell homogenates
showed that both affinity labeled and catalytically active type I enzyme were present on the cytosolic surface of the basolateral
region of the renal cell membrane. |
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Bibliography: | ObjectType-Article-1 SourceType-Scholarly Journals-1 ObjectType-Feature-2 content type line 23 |
ISSN: | 0021-9258 1083-351X |
DOI: | 10.1016/S0021-9258(18)99157-X |