Graduate School of Environmental and Human Sciences, Meijo University, Nagoya 468-8502, Japan.
Graduate School of Environmental and Human Sciences, Meijo University, Nagoya 468-8502, Japan; Research Institute, Meijo University, Nagoya 468-8502, Japan.
Plant Physiol Biochem. 2018 Jun;127:248-255. doi: 10.1016/j.plaphy.2018.03.032. Epub 2018 Mar 30.
Betaine (trimethylglycine) is an important compatible solute that accumulates in response to abiotic stresses such as drought and salinity. Biosynthetic pathways of betaine have been extensively studied, but it remains to be clarified on algae. A diatom Thalassiosira pseudonana CCMP1335 is an important component of marine ecosystems. Here we show that the genome sequence of Thalassiosira suggests the presence of two biosynthetic pathways for betaine, via three step methylation of glycine and via two step oxidation of choline. The choline oxidation via choline dehydrogenase was suggested and its sequential characteristics were analyzed. A candidate gene TpORF1 for glycine methylation encodes a protein consisted of 574 amino acids with two putative tandem repeat methyltransferase domains. The TpORF1 was expressed in E. coli, and the purified protein was shown to synthesize betaine via three step methylation of glycine and designated as TpGSDMT. The proteins containing C-terminal half or N-terminal half were expressed in E. coli and exhibited the methyl transferase activities with different substrate specificity for glycine, sarcosine and dimethylglycine. Upregulation of TpGSDMT transcription and betaine levels were observed at high salinity, suggesting the importance of TpGSDMT for salt tolerance in T. pseudonana cells.
甜菜碱（三甲基甘氨酸）是一种重要的相容溶质，可在干旱和盐度等非生物胁迫下积累。甜菜碱的生物合成途径已得到广泛研究，但在藻类中仍有待阐明。一种硅藻拟菱形藻 CCMP1335 是海洋生态系统的重要组成部分。在这里，我们表明，拟菱形藻的基因组序列表明存在两种甜菜碱生物合成途径，通过甘氨酸的三步甲基化和胆碱的两步氧化。建议通过胆碱脱氢酶进行胆碱氧化，并对其顺序特征进行了分析。甘氨酸甲基化的候选基因 TpORF1 编码由 574 个氨基酸组成的蛋白质，具有两个假定的串联重复甲基转移酶结构域。TpORF1 在大肠杆菌中表达，纯化的蛋白通过甘氨酸的三步甲基化合成甜菜碱，并命名为 TpGSDMT。在大肠杆菌中表达了包含 C 末端一半或 N 末端一半的蛋白质，并表现出对甘氨酸、肌氨酸和二甲氨基甘氨酸具有不同底物特异性的甲基转移酶活性。在高盐度下观察到 TpGSDMT 转录和甜菜碱水平的上调，表明 TpGSDMT 在拟菱形藻细胞的耐盐性中很重要。
