The cloning and characterization of α‐galactosidase present during and following germination of tomato (Lycopersicon esculentum Mill.) seed

• Published in: Journal of experimental botany Vol. 52; no. 359; pp. 1239 - 1249
• Main Authors: Feurtado, J. Allan, Banik, Mitali, Bewley, J. Derek
• Format: Journal Article
• Language: English
• Published: Oxford Oxford University Press 01.06.2001; OXFORD UNIVERSITY PRESS
• Subjects: Agronomy. Soil science and plant productions; alpha-Galactosidase - chemistry; alpha-Galactosidase - classification; alpha-Galactosidase - genetics; Amino Acid Sequence; Biological and medical sciences; Blotting, Northern; Blotting, Southern; Cell and Molecular Biology, Biochemistry and Molecular Physiology; Cell walls; Cloning, Molecular; Complementary DNA; DNA; DNA, Complementary - isolation & purification; DNA, Plant; Economic plant physiology; Embryo development. Germination; Embryos; Endosperm; Enzymes; Fundamental and applied biological sciences. Psychology; Gels; gene expression; Germination; Germination and dormancy; Growth and development; Lycopersicon esculentum - enzymology; Lycopersicon esculentum - genetics; Molecular Sequence Data; Phylogeny; Plant physiology and development; Seeds; Seeds - enzymology; Seeds - genetics; Sequence Homology, Amino Acid; Soybeans; tomato seed; α‐Galactosidase; Embryo; Time variation; Characterization; Dicotyledones; Angiospermae; Lycopersicon esculentum; Endosperm; Developmental stage; Solanaceae; Molecular cloning; Localization; Aminoacid sequence; Leader peptide; Enzyme; Phylogenetic tree; Germination; Tissue specificity; Vegetable crop; α-Galactosidase; Micropyle; Glycosidases; Seed; Hydrolases; Spermatophyta; O-Glycosidases
• ISSN: 0022-0957; 1460-2431
• DOI: 10.1093/jexbot/52.359.1239

α‐Galactosidase (EC 3.2.1.22) is present in the embryo, micropylar and lateral endosperm of seeds of tomato during and following germination. Its activity is unchanged even when germination of the seeds is prevented by an osmoticum. It is also present in the developing and mature dry seed. A cDNA clone for tomato seed α‐galactosidase (LeaGal) has been isolated and the characteristics of the protein deduced; the predicted molecular mass of the mature enzyme is 39.8 kDa, with a pI of 4.91. The tomato α‐galactosidase has a high homology (>62%) at the amino acid level with that of other plant α‐galactosidases. A hydrophobic signal peptide region is identified which is indicative that the enzyme enters the lumen of the endoplasmic reticulum during its translation, prior to its export to the protein body or cell wall, the presumed sites of its substrates. Using amino acid alignment and phylogenetic analysis, key amino acids have been identified, and relationships to other α‐galactosidases inferred. Southern hybridization analyses show that the enzyme is derived from a single gene (for which a partial sequence has been obtained) and yet there are at least three different isoforms within the seed; post‐translational modifications are thus presumed to occur. From Northern hybridization studies it is evident that α‐galactosidase transcripts are present in the lateral and micropylar endosperm during and following germination, and also to a lesser extent in the embryo.