[Functional characterization and enzymatic properties of flavonoid glycosyltransferase gene CtUGT49 in Carthamus tinctorius].

Carthami Flos, as a traditional blood-activating and stasis-resolving drug, possesses anti-tumor, anti-inflammatory, and immunomodulatory pharmacological activities. Flavonoid glycosides are the main bioactive components in Carthamus tinctorius. Glycosyltransferase deserves to be studied in depth as a downstream modification enzyme in the biosynthesis of active glycoside compounds. This study reported a flavonoid glycosyltransferase CtUGT49 from C. tinctorius based on the transcriptome data, followed by bioinformatic analysis and the investigation of enzymatic properties. The open reading frame(ORF) of the gene was 1 416 bp, encoding 471 amino acid residues with the molecular weight of about 52 kDa. Phylogenetic analysis showed that CtUGT49 belonged to the UGT73 family. According to in vitro enzymatic results, CtUGT49 could catalyze naringenin chalcone to the prunin and choerospondin, and catalyze phloretin to phlorizin and trilobatin, exhibiting good substrate versatility. After the recombinant protein CtUGT49 was obtained by hetero-logous expression and purification, the enzymatic properties of CtUGT49 catalyzing the formation of prunin from naringenin chalcone were investigated. The results showed that the optimal pH value for CtUGT49 catalysis was 7.0, the optimal temperature was 37 ℃, and the highest substrate conversion rate was achieved after 8 h of reaction. The results of enzymatic kinetic parameters showed that the K_m value was 209.90 µmol·L~(-1) and k_(cat) was 48.36 s~(-1) calculated with the method of Michaelis-Menten plot. The discovery of the novel glycosyltransferase CtUGT49 is important for enriching the library of glycosylation tool enzymes and provides a basis for analyzing the glycosylation process of flavonoid glycosides in C. tinctorius.

[Seed morphology and germination characteristics of Tripterygium wilfordii].

The seeds of Tripterygium wilfordii are characterized by dormancy and a long germination cycle under natural sowing conditions. In this study, we developed a method for rapid germination of T. wilfordii seeds by analyzing the size, morphology, thousand-grain weight, viability, moisture content, physicochemical properties, and seed germination rates under different germination conditions. The seeds of T. wilfordii were fine columnar with a thick and hard outer seed coat. They had the length of 6.69 mm, the width of 2.14 mm, the thickness of 1.68 mm, the thousand-grain weight of 8.99 g, the moisture content of 8.86%, the soluble sugar content of 21.3 mg·g~(-1), the starch content of 28.9 mg·g~(-1), the soluble protein content of 44.2 mg·g~(-1), and the seed viability of only 54.0%. The seeds were respectively treated with distilled water, ultrasonication, low-temperature storage, 50 ℃ water, 100 mg·L~(-1) 6-BA, 0.6% KMnO_4, 1% KNO_3, 50 mg·L~(-1) NAA, and 100 mg·L~(-1) GA_3 solution. The results showed that soaking the seeds in 100 mg·L~(-1) GA_3 solution significantly promoted the germination. Further, the seeds were soaked in 50, 100, 250, 500, and 1 000 mg·L~(-1) GA_3 solutions, which demonstrated that high concentration(500 mg·L~(-1), 1 000 mg·L~(-1)) of GA_3 solutions increased the germination rate and speed and shortened the germination cycle from more than 3 months to less than 15 days. The findings of this study are of great significance to the breeding of T. wilfordii and lay a foundation for the large-scale propagation of T. wilfordii seeds and the excavation of T. wilfordii germplasm resources.