title: Biochemical characterization of rice trehalose-6-phosphate phosphatases supports distinctive functions of these plant enzymes.
abstract: Substantial levels of trehalose accumulate in bacteria, fungi, and invertebrates, where it serves as a storage carbohydrate or as a protectant against environmental stresses. In higher plants, trehalose is detected at fairly low levels; therefore, a regulatory or signaling function has been proposed for this molecule. In many organisms, trehalose-6-phosphate phosphatase is the enzyme governing the final step of trehalose biosynthesis. Here we report that OsTPP1 and OsTPP2 are the two major trehalose-6-phosphate phosphatase genes expressed in vegetative tissues of rice. Similar to results obtained from our previous OsTPP1 study, complementation analysis of a yeast trehalose-6-phosphate phosphatase mutant and activity measurement of the recombinant protein demonstrated that OsTPP2 encodes a functional trehalose-6-phosphate phosphatase enzyme. OsTPP2 expression is transiently induced in response to chilling and other abiotic stresses. Enzymatic characterization of recombinant OsTPP1 and OsTPP2 revealed stringent substrate specificity for trehalose 6-phosphate and about 10 times lower K(m) values for trehalose 6-phosphate as compared with trehalose-6-phosphate phosphatase enzymes from microorganisms. OsTPP1 and OsTPP2 also clearly contrasted with microbial enzymes, in that they are generally unstable, almost completely losing activity when subjected to heat treatment at 50 degrees C for 4 min. These characteristics of rice trehalose-6-phosphate phosphatase enzymes are consistent with very low cellular substrate concentration and tightly regulated gene expression. These data also support a plant-specific function of trehalose biosynthesis in response to environmental stresses.
authors.affiliation: Crop Cold Tolerance Research Team, National Agricultural Research Center for Hokkaido Region, National Agriculture and Food Research Organization, Hitsujigaoka 1, Toyohira-ku, Sapporo 062-8555, Japan.
authors.forename: Shuhei
authors.initials: S
authors.lastname: Shima
authors.forename: Hirokazu
authors.initials: H
authors.lastname: Matsui
authors.forename: Satoshi
authors.initials: S
authors.lastname: Tahara
authors.forename: Ryozo
authors.initials: R
authors.lastname: Imai
mesh_terms: D000595:Amino Acid Sequence; D004795:Enzyme Stability; D015971:Gene Expression Regulation, Enzymologic; D018506:Gene Expression Regulation, Plant; D017343:Genes, Plant; D005816:Genetic Complementation Test; D005982:Glutathione Transferase / Q000378:metabolism; D006358:Hot Temperature; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008969:Molecular Sequence Data; D009154:Mutation; D012275:Oryza / Q000201:enzymology* / Q000235:genetics*; D010744:Phosphoric Monoester Hydrolases / Q000032:analysis / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D010940:Plant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D018517:Plant Roots / Q000201:enzymology; D018520:Plant Shoots / Q000201:enzymology; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000201:enzymology / Q000235:genetics; D036226:Seedlings / Q000201:enzymology; D017386:Sequence Homology, Amino Acid; D013379:Substrate Specificity
chemical_list: D010940:Plant Proteins; D011993:Recombinant Fusion Proteins; D005982:Glutathione Transferase; C023178:trehalose-phosphatase; D010744:Phosphoric Monoester Hydrolases