Abstract
9-O-Acetylation of sialic acid is known as a cell type-specific modification of secretory and plasma membrane glycoconjugates of higher vertebrates with important functions in modulating cell-cell recognition. Using a recombinant probe derived from influenza C virus hemagglutinin, we discovered 9-O-acetylated protein in the Golgi complex of various cell lines, most of which did not display 9-O-acetylated sialic acid on the cell surface. All cell lines expressed a sulfated glycoprotein of 50 kDa (sgp50) carrying 9-O-acetylated sialic acids, which was used as a model substrate. Like gp40, the major receptor for influenza C virus of Madin-Darby canine kidney I cells, sgp50 is 9-O-acetylated on O-linked glycans. However, gp40 was not 9-O-acetylated when expressed in Madin-Darby canine kidney II or COS-7 cells. The results demonstrate the existence of two 9-O-acetylation machineries for O-glycosylated proteins with distinct substrate specificities. The widespread occurrence of 9-O-acetylated protein in the Golgi furthermore suggests an additional intracellular role for this modification.
Highlights
O-Acetyl transferases for sialic acids have resisted purification or cloning far
We discovered that all cell lines tested, including some that were previously shown to be resistant to infection by influenza C virus and considered to lack 9-O-acetyl sialic acid, stained positive intracellularly in the Golgi apparatus
Intracellular 9-O-Acetylated Sialic Acid Detected with CHEFcD Fluorescence Microscopy—CHE-FcD/protein ASepharose (CHE)-Fc, the fusion protein of influenza C virus hemagglutinin-esterase with the constant region of human IgG, efficiently binds to 9-O-acetylated sialic acid if the esterase is inactivated by diisopropyl fluorophosphate treatment [20]
Summary
O-Acetyl transferases for sialic acids have resisted purification or cloning far. All cell lines expressed a sulfated glycoprotein of 50 kDa (sgp50) carrying 9-O-acetylated sialic acids, which was used as a model substrate.
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