Heterologous biomimetic synthesis of active ingredients in traditional Chinese medicine:a new mode for protection and development of traditional Chinese medicine resources / 中国中药杂志

Heterologous biomimetic synthesis of the active ingredients of traditional Chinese medicine(TCM) is a new mode of resource acquisition and has shown great potential in the protection and development of TCM resources. According to synthetic biology and by constructing biomimetic microbial cells and imitating the synthesis of active ingredients in medicinal plants and animals, the key enzymes obtained from medicinal plants and animals are scientifically designed and systematically reconstructed and optimized to realize the heterologous synthesis of the active ingredients in microorganisms. This method ensures an efficient and green acquisition of target products, and also achieves large-scale industrial production, which is conducive to the production of scarce TCM resources. Additiona-lly, the method playes a role in agricultural industrialization, and provides a new option for promoting the green and sustainable deve-lopment of TCM resources. This review systematically summarized the important progress in the heterologous biomimetic synthesis of TCM active ingredients from three research areas: biosynthesis of terpenoids, flavonoids, phenylpropanoids, alkaloids and other active ingredients, key points and difficulties in heterologous biomimetic synthesis, and biomimetic cells with complex TCM ingredients. This study facilitated the application of new generation of biotechnology and theory to the development of TCM.

Construction of cell factories for high production of ginsenoside Rh_2 in Saccharomyces cerevisiae / 中国中药杂志

Ginsenoside Rh_2 is a rare active ingredient in precious Chinese medicinal materials such as Ginseng Radix et Rhizoma, Notoginseng Radix et Rhizoma, and Panacis Quinquefolii Radix. It has important pharmacological activities such as anti-cancer and improving human immunity. However, due to the extremely low content of ginsenoside Rh_2 in the source plants, the traditional way of obtaining it has limitations. This study intended to apply synthetic biological technology to develop a cell factory of Saccharomyces cerevisiae to produce Rh_2 by low-cost fermentation. First, we used the high protopanaxadiol(PPD)-yielding strain LPTA as the chassis strain, and inserted the Panax notoginseng enzyme gene Pn1-31, together with yeast UDP-glucose supply module genes[phosphoglucose mutase 1(PGM1), α-phosphoglucose mutase(PGM2), and uridine diphosphate glucose pyrophosphorylase(UGP1)], into the EGH1 locus of yeast chromosome. The engineered strain LPTA-RH2 produced 17.10 mg·g~(-1) ginsenoside Rh_2. This strain had low yield of Rh_2 while accumulated much precursor PPD, which severely restricted the application of this strain. In order to further improve the production of ginsenoside Rh_2, we strengthened the UDP glucose supply module and ginsenoside Rh_2 synthesis module by engineered strain LPTA-RH2-T. The shaking flask yield of ginsenoside Rh_2 was increased to 36.26 mg·g~(-1), which accounted for 3.63% of the dry weight of yeast cells. Compared with those of the original strain LPTA-RH2, the final production and the conversion efficiency of Rh_2 increased by 112.11% and 65.14%, respectively. This study provides an important basis for further obtaining the industrial-grade cell factory for the production of ginsenoside Rh_2.

Study of heterologous efficient synthesis of cucurbitadienol / 中国中药杂志

Cucurbitadienol has anti-inflammation, anti-cancer activities, and acts as a precursor of traditional Chinese medicine active ingredients mogroside and cucurbitacine. For construction of a Sacchromyces cerevisiae cell factory for production of cucurbitadienol, we firstly cloned a cucurbitadienol synthase (CBS) gene from Siraitia grosvenorii. Then, through heterologous expression of CBS in the triterpenoid chassis strain WD-2091, the engineered strain could produced 27.44 mg•L ⁻¹ cucurbitadienol, which was determined by GC-MS. Further regulation of CBS expression led to cucurbitadienol's titer increasing by 202.07% and reaching 82.89 mg•L ⁻¹ in the shake flask fermentation and 1 724.10 mg•L ⁻¹ in the high cell density fermentation. Our research promotes the cucurbitane-type tetracyclic triterpenoids synthesis pathway analysis progress and provides the basis for further obtaining cell factories for production of cucurbitadienol tetracyclic triterpenoids.

Research progress of synthetic biology for tanshinones / 中国中药杂志

Synthetic biology research methods which design and build a new artificial biological systems (medicinal plants or microorganisms system) with specific physiological functions through clarifying and simulating the basic law of the biosynthesis of active components of traditional Chinese medicine, is considered to be a potential method to produce an abundant resources of bioactive components. Tanshinones is a kind of diterpene quinone compounds with important pharmacological activities from traditional Chinese medicine Salvia miltiorrhiza. This article systematically introduced the research progress of the synthetic biology of S. miltiorrhiza, in order to provide references for studies on other terpenoid bioactive components of traditional Chinese medicines, and give new research strategies for the sustainable development of traditional Chinese medicine resources.

Status and future of natural resource for Chinese materia medica / 中国中药杂志

For thousands of years, the natural resource for Chinese materiamedica has been the foundation of the traditional Chinese medicine industry, which provides abundant medicine for human. In recent years, increasing demands and irrational exploitation led to a lot of problems such as rapid decrease of traditional Chinese herbs reserves, low quality of medicine and dismishing traditional cultures. These restricted the development of the traditional Chinese medicine. To solve these problems, scientists have done much work on investigating traditional Chinese medicine resources, exploring the metabolic pathway of bioactive ingredients, cultivating new varieties, and carrying out synthetic biology. These studies provided a theoretical basis for sustainable utilizationand future developmentof traditional Chinese medicine resources.

Cloning and characterization of cDNA encoding Psammosilene tunicoides squalene synthase / 药学学报

The total triterpene saponins of Psammosilene tunicoides have significant pharmacologic activity. Psammosilene tunicoides squalene synthase (PSS) is a gateway enzyme to regulate the biosynthesis of total triterpene saponins extracted from the root of Psammosilene tunicoides which is an endangered species. In this paper, cDNA encoding of PSS was cloned by the degenerate primer PCR and rapid-amplification of cDNA ends (RACE). The full-length of cDNA of PSS is 1663 bp, with an open reading frame (ORF) of 1 245 bp, encoding 414 amino acid polypeptide (calculated molecular mass, 47.69 kDa), 5'UTR (untranslated region) and 3'UTR are 260 bp and 158 bp, respectively. The deduced amino acid sequence of PSS has higher homology with the known squalene synthases of several species such as Panax notoginseng (83%), Panax ginseng (82%) and Glycyrrhiza glabra (82%) than that with Schizosacharomyces pombe (35%), Candida albicans (39%) and Homo sapiens (47%). The characterization of PSS was done by a series of methods, such as prokaryotic expression, the activity of enzyme in vitro, capillary gas chromatography (GC) and capillary gas chromatography mass spectrometry (GC-MS). The results showed that the cell-free extract of E. coli transformed with the recombinant plasmid can effectively convert farnesyl diphosphate into squalene in vitro. GenBank accession number is EF585250. Our research provided important base for the study of Psammosilene tunicoides secondary metabolism and metabolic engineering.