Cloning and characterization of genes encoding trehalose-6-phosphate synthase (TPS1) and trehalose-6-phosphate phosphatase (TPS2) from Zygosaccharomyces rouxii.
In many organisms, trehalose protects against several environmental stresses, such as heat, desiccation, and salt, probably by stabilizing protein structures and lipid membranes. Trehalose synthesis in yeast is mediated by a complex of trehalose-6-phosphate synthase (TPS1) and trehalose-6-phosphate phosphatase (TPS2). In this study, genes encoding TPS1 and TPS2 were isolated from Zygosaccharomyces rouxii (designated ZrTPS1 and ZrTPS2, respectively). They were functionally identified by their complementation of the tps1 and tps2 yeast deletion mutants, which are unable to grow on glucose medium and with heat, respectively. Full-length ZrTPS1 cDNA is composed of 1476 nucleotides encoding a protein of 492 amino acids with a molecular mass of 56 kDa. ZrTPS2 cDNA consists of 2843 nucleotides with an open reading frame of 2700 bp, which encodes a polypeptide of 900 amino acids with a molecular mass of 104 kDa. The amino acid sequence encoded by ZrTPS1 has relatively high homology with TPS1 of Saccharomyces cerevisiae and Schizosaccharomyces pombe, compared with TPS2. Western blot analysis showed that the antibody against S. cerevisiae TPS1 recognizes ZrTPS1. Under normal growth conditions, ZrTPS1 and ZrTPS2 were highly and constitutively expressed, unlike S. cerevisiae TPS1 and TPS2. Salt stress and heat stress reduced the expression of the ZrTPS1 and ZrTPS2 genes, respectively.
Kwon
Hawk-Bin
HB
Division of Applied Biological Sciences, Sunmoon University, 336-708, Asan, South Korea.
Yeo
Eun-Taeg
ET
Hahn
Sang-Eun
SE
Bae
Shin-Chul
SC
Kim
Dool-Yi
DY
Byun
Myung-Ok
MO
D000595:Amino Acid Sequence; D001483:Base Sequence; D015152:Blotting, Northern; D015153:Blotting, Western; D003001:Cloning, Molecular; D003080:Cold Temperature; D005656:Fungal Proteins / Q000378:metabolism; D015966:Gene Expression Regulation, Fungal; D005816:Genetic Complementation Test; D005964:Glucosyltransferases / Q000096:biosynthesis / Q000235:genetics*; D006358:Hot Temperature; D008969:Molecular Sequence Data; D010744:Phosphoric Monoester Hydrolases / Q000096:biosynthesis / Q000235:genetics*; D012333:RNA, Messenger / Q000096:biosynthesis / Q000235:genetics; D019105:Random Amplified Polymorphic DNA Technique; D020133:Reverse Transcriptase Polymerase Chain Reaction; D017386:Sequence Homology, Amino Acid; D012965:Sodium Chloride / Q000378:metabolism; D020068:Zygosaccharomyces / Q000201:enzymology* / Q000235:genetics*
D005656:Fungal Proteins; D012333:RNA, Messenger; D012965:Sodium Chloride; D005964:Glucosyltransferases; C023180:trehalose-6-phosphate synthase; C023178:trehalose-phosphatase; D010744:Phosphoric Monoester Hydrolases