A Novel Alliinase from Onion Roots. Biochemical Characterization and cDNA Cloning

• Published in: Plant physiology (Bethesda) 2000-04, Vol.122 (4), p.1269-1279
• Main Authors: Jane E. Lancaster, Martin L. Shaw, Meeghan D. Pither Joyce, John A. Mc Callum, McManus, Michael T.
• Format: Article
• Language: English
• Subjects: Agronomy. Soil science and plant productions; Amino Acid Sequence; Amino acids; Base Sequence; Biological and medical sciences; Biotechnology; Carbon-Sulfur Lyases - genetics; Carbon-Sulfur Lyases - metabolism; Cloning; Cloning, Molecular; Complementary DNA; DNA Primers; DNA, Complementary; Economic plant physiology; Enzymes; Fundamental and applied biological sciences. Psychology; Garlic; Gels; Lectins; Liquid chromatography; Metabolism; Molecular Sequence Data; Nutrition. Photosynthesis. Respiration. Metabolism; Onions; Onions - enzymology; Plant physiology and development; Plant Roots - enzymology; Protein isoforms; Proteins; RNA; Roots; Sequence Homology, Amino Acid; Sulfoxides
• ISSN: 0032-0889; 1532-2548
• DOI: 10.1104/pp.122.4.1269

We have purified a novel alliinase (EC 4.4.1.4) from roots of onion (Allium cepa L.). Two isoforms with alliinase activity (I and II) were separated by concanavalin A-Sepharose and had molecular masses of 52.7 (I) and 50.5 (II) kD on sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and 51 (I) and 57.5 (II) kD by gel filtration fast-protein liquid chromatography. Isoform I had an isoelectric point of 9.3, while isoform II had isoelectric points of 7.6, 7.9, 8.1, and 8.3. The isoforms differed in their glycosylation. Both contained xylose/fucose containing complex-type N-linked glycans, and isoform II also contained terminal mannose structures. Both isoforms had activity with S-alk(en)yl-L-cysteine sulfoxides. Unlike other allium alliinases, A. cepa root isoforms had cystine lyase activity. We cloned a gene from A. cepa root cDNA and show that it codes for A. cepa root alliinase protein. Homology to other reported allium alliinase genes is 50%. The gene coded for a protein of mass 51.2 kD, with two regions of deduced amino acid sequence identical to a 25- and a 40-amino acid region, as determined experimentally. The A. cepa root alliinase cDNA was expressed mainly in A. cepa roots. The structure and function of the alliinase gene family is discussed.