Cloning and expression pattern of a gene encoding an α-xylosidase active against xyloglucan oligosaccharides from Arabidopsis

An alpha -xylosidase active against xyloglucan oligosaccharides was purified from cabbage (Brassica oleracea var. capitata) leaves. Two peptide sequences were obtained from this protein, the N-terminal and an internal one, and these were used to identify an Arabidopsis gene coding for an alpha -xylosidase that we propose to call AtXYL1. it has been mapped to a region of chromosome I between markers at 100.44 and 107.48 cM. AtXYL1 comprised three exons and encoded a peptide that was 915 amino acids long, with a potential signal peptide of 22 amino acids and eight possible N-glycosylation sites. The protein encoded by AtXYL1 showed the signature regions of family 31 glycosyl hydrolases, which comprises not only alpha -xylosidases, but also alpha -glucosidases. The alpha -xylosidase activity is present in apoplastic extractions from Arabidopsis seedlings, as suggested by the deduced signal peptide. The first eight levels from Arabidopsis plants were harvested to analyze alpha -xylosidase activity and AtXYL1 expression levels. Both increased from older to younger leaves, where xyloglucan turnover is expected to be higher. When this gene was introduced in a suitable expression vector and used to transform Saccharomyces cerevisiae, significantly higher alpha -xylosidase activity was detected in the yeast cells. cu-Glucosidase activity was also increased in the transformed cells, although to a lesser extent. These results show that AtXYL1 encodes for an apoplastic alpha -xylosidase active against xyloglucan oligosaccharides that probably also has activity against p-nitrophenyl-alpha -D-glucoside.