Abstract
Art v 1, the major allergen of mugwort (Artemisia vulgaris) pollen contains galactose and arabinose. As the sera of some allergic patients react with natural but not with recombinant Art v 1 produced in bacteria, the glycosylation of Art v 1 may play a role in IgE binding and human allergic reactions. Chemical and enzymatic degradation, mass spectrometry, and 800 MHz (1)H and (13)C nuclear magnetic resonance spectroscopy indicated the proline-rich domain to be glycosylated in two ways. We found a large hydroxyproline-linked arabinogalactan composed of a short beta1,6-galactan core, which is substituted by a variable number (5-28) of alpha-arabinofuranose residues, which form branched side chains with 5-, 2,5-, 3,5-, and 2,3,5-substituted arabinoses. Thus, the design of the Art v 1 polysaccharide differs from that of the well known type II arabinogalactans, and we suggest it be named type III arabinogalactan. The other type of glycosylation was formed by single (but adjacent) beta-arabinofuranoses linked to hydroxyproline. In contrast to the arabinosylation of Ser-Hyp(4) motifs in other hydroxyproline-rich glycoproteins, such as extensins or solanaceous lectins, no oligo-arabinosides were found in Art v 1. Art v 1 and parts thereof produced by alkaline degradation, chemical deglycosylation, proteolytic degradation, and/or digestion with alpha-arabinofuranosidase were used in enzyme-linked immunosorbent assay and immunoblot experiments with rabbit serum and with the sera of patients. Although we could not observe antibody binding by the polysaccharide, the single hydroxyproline-linked beta-arabinose residues appeared to react with the antibodies. Mono-beta-arabinosylated hydroxyproline residues thus constitute a new, potentially cross-reactive, carbohydrate determinant in plant proteins.
Highlights
Art v 1, the major allergen from the pollen of mugwort (Artemisia vulgaris), which constitutes a prominent allergen in the temperate climate zone, has recently been characterized and sequenced, and its cDNA has been cloned (1)
We found a large hydroxyproline-linked arabinogalactan composed of a short 1,6-galactan core, which is substituted by a variable number (5–28) of ␣-arabinofuranose residues, which form branched side chains with 5, 2,5, 3,5, and 2,3,5-substituted arabinoses
In the present study we report on the structure of the arabinogalactan polysaccharide and on the unexpected finding of a new type of O-glycosylation of the mugwort (A. vulgaris) pollen allergen Art v 1 as well as on the antibody binding abilities of these posttranslational modifications
Summary
Art v 1, the major allergen from the pollen of mugwort (Artemisia vulgaris), which constitutes a prominent allergen in the temperate climate zone, has recently been characterized and sequenced, and its cDNA has been cloned (1). The carbohydrate part of AGPs consists of a hydroxyproline-linked 1,3-D-galactan backbone having 1,6-Dgalactan side chains or kinks, which in turn are substituted by arabinoses and other less abundant sugar residues (8 –10) These type II arabinogalactans (in contrast to the type I with 1,4 linkages (8)) typically contain more galactose than arabinose. Extensins and solanaceous lectins exhibit two different types of O-glycosylation, single ␣-galactosyl residues linked to Ser residues (11, 12) and short arabinan chains linked directly to hydroxyproline (5, 8, 10, 13–15) Such arabins have been reported to consist of 1–5 arabinofuranose residues, which are joined by 1,2 and 1,3 linkages (13, 15–17). In the present study we report on the structure of the arabinogalactan polysaccharide and on the unexpected finding of a new type of O-glycosylation of the mugwort (A. vulgaris) pollen allergen Art v 1 as well as on the antibody binding abilities of these posttranslational modifications
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