BIOL 112 |
| The Golgi-resident enzyme N-acetylglucosamine-6-sulfotransferase-1 (GlcNAc6ST-1) is responsible for sulfation of GlcNAc on the L-selectin ligand. GlcNAc6ST-1 is a glycosylated type II transmembrane protein. Here we report the sites at which GlcNAc6ST-1 is modified with N-linked glycans and the effects of glycosylation on enzyme activity. We prepared six GlcNAc6ST-1 mutants in which putative N-linked sites were absent and analyzed these mutants in cellular and in vitro assays. Western blotting revealed two N-linked glycans on GlcNAc6ST-1; mutations reduced the number of glycans. Cellular activity of the mutants was measured by flow cytometry, indicating that the absence of ¬N¬-linked glycans can diminish or abrogate enzyme activity. Subcellular localization of GlcNAc6ST-1 was generally unaffected by mutations: immunofluorescence microscopy was used to show that most mutants localize to the Golgi, like the wild-type enzyme. A notable exception is the N428A mutant, which lacks cellular activity and exhibits altered localization. |
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Chemistry and Metabolism
4:30 PM-6:30 PM, Tuesday, 12 September 2006 Moscone Center -- Hall D, Poster
Sci-Mix
Division of Biological Chemistry |