Yin Qinggang, Zhang Jing, Wang Shuhui, Cheng Jintang, Gao Han, Guo Cong, Ma Lianbao, Sun Limin, Han Xiaoyan, Chen Shilin, Liu An
Key Laboratory of Beijing for Identification and Safety Evaluation of Chinese Medicine, Institute of Chinese Materia Medica, China Academy of Chinese Medical Sciences, Beijing, 100700, China.
Artemisinin Research Center, China Academy of Chinese Medical Sciences, Beijing, 100700, China.
Hortic Res. 2021 Nov 1;8(1):229. doi: 10.1038/s41438-021-00658-0.
As auxins are among the most important phytohormones, the regulation of auxin homeostasis is complex. Generally, auxin conjugates, especially IAA glucosides, are predominant at high auxin levels. Previous research on terminal glucosylation focused mainly on the O-position, while IAA-N-glucoside and IAA-Asp-N-glucoside have been neglected since their discovery in 2001. In our study, IAA-Asp-N-glucoside was found to be specifically abundant (as high as 4.13 mg/g) in the seeds of 58 ginkgo cultivars. Furthermore, a novel N-glucosyltransferase, termed GbNGT1, was identified via differential transcriptome analysis and in vitro enzymatic testing. It was found that GbNGT1 could catalyze IAA-Asp and IAA to form their corresponding N-glucosides. The enzyme was demonstrated to possess a specific catalytic capacity toward the N-position of the IAA-amino acid or IAA from 52 substrates. Docking and site-directed mutagenesis of this enzyme confirmed that the E15G mutant could almost completely abolish its N-glucosylation ability toward IAA-Asp and IAA in vitro and in vivo. The IAA modification of GbNGT1 and GbGH3.5 was verified by transient expression assay in Nicotiana benthamiana. The effect of GbNGT1 on IAA distribution promotes root growth in Arabidopsis thaliana.
由于生长素是最重要的植物激素之一，生长素稳态的调节十分复杂。一般来说，生长素共轭物，尤其是吲哚-3-乙酸（IAA）糖苷，在高生长素水平时占主导地位。以往关于末端糖基化的研究主要集中在O-位，而IAA-N-葡萄糖苷和IAA-天冬酰胺-N-葡萄糖苷自2001年被发现以来一直被忽视。在我们的研究中，发现IAA-天冬酰胺-N-葡萄糖苷在58个银杏品种的种子中特别丰富（高达4.13毫克/克）。此外，通过差异转录组分析和体外酶活性测试，鉴定出一种新的N-葡萄糖基转移酶，命名为GbNGT1。发现GbNGT1可以催化IAA-天冬酰胺和IAA形成它们相应的N-葡萄糖苷。该酶对52种底物中的IAA-氨基酸或IAA的N-位具有特异性催化能力。该酶的对接和定点诱变证实，E15G突变体在体外和体内几乎可以完全消除其对IAA-天冬酰胺和IAA的N-糖基化能力。通过在本氏烟草中的瞬时表达试验验证了GbNGT1和GbGH3.5对IAA的修饰作用。GbNGT1对IAA分布的影响促进了拟南芥根系的生长。
