Arabidopsis thaliana VTC4 Encodes L-Galactose-1-P Phosphatase, a Plant Ascorbic Acid Biosynthetic Enzyme
| Autor: | Nicholas Smirnoff, Marjorie J. Raymond, Patricia L. Conklin, M.N. Isupov, John Dowdle, Jennifer A. Littlechild, Stephan Gatzek, Susanne Rolinski, Glen L. Wheeler |
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| Rok vydání: | 2006 |
| Předmět: |
Molecular Sequence Data
Phosphatase Mutant Arabidopsis Ascorbic Acid Biology Genes Plant Biochemistry Gene product chemistry.chemical_compound Biosynthesis Animals Humans Amino Acid Sequence Molecular Biology Phylogeny DNA Primers chemistry.chemical_classification Base Sequence Sequence Homology Amino Acid Reverse Transcriptase Polymerase Chain Reaction Wild type Galactose Cell Biology biology.organism_classification Ascorbic acid Phosphoric Monoester Hydrolases Enzyme chemistry Mutation Mannose |
| Zdroj: | Journal of Biological Chemistry. 281:15662-15670 |
| ISSN: | 0021-9258 |
| DOI: | 10.1074/jbc.m601409200 |
| Popis: | In plants, a proposed ascorbate (vitamin C) biosynthesis pathway occurs via GDP-D-mannose (GDP-D-Man), GDP-L-galactose (GDP-L-Gal), and L-galactose. However, the steps involved in the synthesis of L-Gal from GDP-L-Gal in planta are not fully characterized. Here we present evidence for an in vivo role for L-Gal-1-P phosphatase in plant ascorbate biosynthesis. We have characterized a low ascorbate mutant (vtc4-1) of Arabidopsis thaliana, which exhibits decreased ascorbate biosynthesis. Genetic mapping and sequencing of the VTC4 locus identified a mutation (P92L) in a gene with predicted L-Gal-1-P phosphatase activity (At3g02870). Pro-92 is within a beta-bulge that is conserved in related myo-inositol monophosphatases. The mutation is predicted to disrupt the positioning of catalytic amino acid residues within the active site. Accordingly, L-Gal-1-P phosphatase activity in vtc4-1 was approximately 50% of wild-type plants. In addition, vtc4-1 plants incorporate significantly more radiolabel from [2-(3)H]Man into L-galactosyl residues suggesting that the mutation increases the availability of GDP-L-Gal for polysaccharide synthesis. Finally, a homozygous T-DNA insertion line, which lacks a functional At3g02870 gene product, is also ascorbate-deficient (50% of wild type) and deficient in L-Gal-1-P phosphatase activity. Genetic complementation tests revealed that the insertion mutant and VTC4-1 are alleles of the same genetic locus. The significantly lower ascorbate and perturbed L-Gal metabolism in vtc4-1 and the T-DNA insertion mutant indicate that L-Gal-1-P phosphatase plays a role in plant ascorbate biosynthesis. The presence of ascorbate in the T-DNA insertion mutant suggests there is a bypass to this enzyme or that other pathways also contribute to ascorbate biosynthesis. |
| Databáze: | OpenAIRE |
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