Although plant secondary metabolites are important source of new drugs, obtaining these compounds is challenging due to their high structural diversity and low abundance. The roots of Astragalus membranaceus are a popular herbal medicine worldwide. It contains a series of cycloartane-type saponins (astragalosides) as hepatoprotective and antivirus components. However, astragalosides exhibit complex sugar substitution patterns which hindered their purification and bioactivity investigation. In this work, glycosyltransferases (GT) from A. membranaceus were studied to synthesize structurally diverse astragalosides. Three new GTs, AmGT1/5 and AmGT9, were characterized as 3-O-glycosyltransferase and 25-Oglycosyltransferase of cycloastragenol respectively. AmGT1 G146V/I variants were obtained as specific 3-O-xylosyltransferases by sequence alignment, molecular modelling and site-directed mutagenesis. A combinatorial synthesis system was established using AmGT1/5/9, AmGT1 G146V/S and the reported AmGT8 and AmGT8 A394F . The system allowed the synthesis of 13 astragalosides in Astragalus root with conversion rates from 22.6% to 98.7%, covering most of the sugar-substitution patterns for astragalosides. In addition, AmGT1 exhibited remarkable sugar donor promiscuity to use 10 different donors, and was used to synthesize three novel astragalosides and ginsenosides. Glycosylation remarkably improved the hepatoprotective and SARS-CoV-2 inhibition activities for triterpenoids. This is one of the first attempts to produce a series of herbal constituents via combinatorial synthesis. The results provided new biocatalytic tools for saponin biosynthesis.
Accurate metabolite characterization
plays a vital role in targeted
metabolomics. Nonetheless, the library of metabolites is still limited,
especially for downstream conjugates, and it is time-consuming to
synthesize each of these compounds due to high structural diversity.
Herein, a green and smart strategy was developed to expand the scope
of targeted metabolomics. The reference standards were synthesized
in a one-pot microscale reaction, and the analytical method was tailored
using the synthetic products. A group of new metabolites, namely bile
acid–amino acid conjugates (BA–AAs), was studied as
a proof-of-concept. First, in total 160 BA–AAs were synthesized
using a small amount (2 mg each) of bile acids and low-toxic reagents
within 4 h. Then, an ultra-high-performance liquid chromatography
/Orbitrap-MS method was established to comprehensively profile 202
bile acid derivatives in 20 min. Finally, the method was applied to
mice with inflammatory bowel disease (IBD) to discover the accumulation
of 70 rare BA–AAs in small intestine and liver, where 55 were
first reported from biosamples. These BA–AAs are farnesoid
X receptor modulators and might contribute to the development of IBD.
Our study demonstrated a feasible approach for the broad-spectrum
targeted metabolomics of bile acids.
Three leaves written in Brāhmī script and kept in the Dunhuang Research Academy turn out to be parts of a bilingual text of Dharmaśarīrasūtra in Sanskrit and Uighur. After analysing several versions of Dharmaśarīrasūtra, it can be inferred that these three fragments belong to the Northern Brāhmī recensions which were circulated along the Northern Silk Road and are different from the Southern Brāhmī recensions popular along the Southern Route, such as the Khotanese version. This paper attempts to transcribe these fragments and make a thorough research on Dharmaśarīrasūtra, taking five relevant Chinese versions into account.
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