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Molecular cloning and characterization of a plant α1,3/4-fucosidase based on sequence tags from almond fucosidase I
- Source :
- Phytochemistry. 67:641-648
- Publication Year :
- 2006
- Publisher :
- Elsevier BV, 2006.
-
Abstract
- Our work with almond peptide N-glycosidase A made us interested also in the alpha1,3/4-fucosidase which is used as a specific reagent for glycoconjugate analysis. The enzyme was purified to presumed homogeneity by a series of chromatographic steps including dye affinity and fast-performance anion exchange chromatography. The 63 kDa band was analyzed by tandem mass spectrometry which yielded several partial sequences. A homology search retrieved the hypothetical protein Q8GW72 from Arabidopsis thaliana. This protein has recently been described as being specific for alpha1,2-linkages. However, cDNA cloning and expression in Pichia pastoris of the A. thaliana fucosidase showed that it hydrolyzed fucose in 3- and 4-linkage to GlcNAc in Lewis determinants whereas neither 2-linked fucose nor fucose in 3-linkage to the innermost GlcNAc residue were attacked. This first cloning of a plant alpha1,3/4-fucosidase also confirmed the identity of the purified almond enzyme and thus settles the notorious uncertainty about its molecular mass. The alpha1,3/4-fucosidase from Arabidopsis exhibited striking sequence similarity with an enzyme of similar substrate specificity from Streptomyces sp. (Q9Z4I9) and with putative proteins from rice.
- Subjects :
- Spectrometry, Mass, Electrospray Ionization
DNA, Complementary
Molecular Sequence Data
Hypothetical protein
Arabidopsis
Sequence alignment
Plant Science
Horticulture
Molecular cloning
Biochemistry
Pichia
Fucose
Acetylglucosamine
Substrate Specificity
Pichia pastoris
chemistry.chemical_compound
Lewis Blood Group Antigens
Complementary DNA
Fucosidase
Cloning, Molecular
Molecular Biology
alpha-L-Fucosidase
biology
Molecular mass
Hydrolysis
food and beverages
Oryza
General Medicine
biology.organism_classification
Molecular biology
Streptomyces
Molecular Weight
Carbohydrate Sequence
chemistry
biology.protein
Prunus
Sequence Alignment
Subjects
Details
- ISSN :
- 00319422
- Volume :
- 67
- Database :
- OpenAIRE
- Journal :
- Phytochemistry
- Accession number :
- edsair.doi.dedup.....373793a1e9546189d51dc559f55be5e0