Purification, characterization, and cell wall localization of an alpha-fucosidase that inactivates a xyloglucan oligosaccharin.
An alpha-fucosidase that releases fucosyl residues from oligosaccharide fragments of xyloglucan, a plant cell wall hemicellulosic polysaccharide, was purified to homogeneity from pea (Pisum sativum) epicotyls using a combination of cation exchange chromatography and isoelectric focusing. The alpha-fucosidase has a molecular mass of 20 kDa according to sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The alpha-fucosidase has an isoelectric point of 5.5. The substrate specificity of the alpha-fucosidase was determined by high performance anion exchange chromatographic analysis of oligosaccharide substrates and products. The enzyme hydrolyzes the terminal alpha-1,2-fucosidic linkage of oligosaccharides and does not cleave p-nitrophenyl-alpha-L-fucoside. The enzyme does not release measurable amounts of fucosyl residues from large polysaccharides. The subcellular localization of alpha-fucosidase in pea stems and leaves has been studied by immunogold cytochemistry. The alpha-fucosidase accumulates in primary cell walls and is not detectable in the middle lamella or in the cytoplasm of 8-day-old stem tissue and 14-day-old leaf tissue. alpha-Fucosidase activity was readily detected in extracts of 8-day-old stem tissue. No significant alpha-fucosidase activity or immunogold labeling of the alpha-fucosidase was detected in 2- and 4-day-old stem tissue indicating that production of alpha-fucosidase is developmentally regulated.
Augur
C
C
Complex Carbohydrate Research Center, University of Georgia, Athens 30602.
Benhamou
N
N
Darvill
A
A
Albersheim
P
P
D000918:Antibody Specificity; D002240:Carbohydrate Sequence; D002473:Cell Wall / Q000201:enzymology* / Q000378:metabolism; D007887:Fabaceae / Q000201:enzymology* / Q000378:metabolism / Q000648:ultrastructure; D005936:Glucans; D007150:Immunohistochemistry; D008969:Molecular Sequence Data; D009844:Oligosaccharides / Q000302:isolation & purification / Q000378:metabolism*; D010946:Plants, Medicinal; D011134:Polysaccharides / Q000378:metabolism*; D013379:Substrate Specificity; D014990:Xylans; D005644:alpha-L-Fucosidase / Q000276:immunology / Q000302:isolation & purification / Q000378:metabolism*
D005936:Glucans; D009844:Oligosaccharides; D011134:Polysaccharides; D014990:Xylans; C029353:xyloglucan; D005644:alpha-L-Fucosidase