scholarly journals GuUGT, a glycosyltransferase from Glycyrrhiza uralensis , exhibits glycyrrhetinic acid 3- and 30-O-glycosylation

2019 ◽  
Vol 6 (10) ◽  
pp. 191121 ◽  
Author(s):  
Ying Huang ◽  
Da Li ◽  
Jinhe Wang ◽  
Yi Cai ◽  
Zhubo Dai ◽  
...  
Keyword(s):  
Biosynthetic Pathway ◽  
Glycyrrhetinic Acid ◽  
Glycyrrhiza Uralensis ◽  
Transcriptome Data ◽  
Bioactive Components ◽  
Triterpenoid Saponins ◽  
Genes Encoding ◽  
Engineered Yeast ◽  
Biochemical Analyses

Glycyrrhiza uralensis is a well-known herbal medicine that contains triterpenoid saponins as the predominant bioactive components, and these compounds include glycyrrhetinic acid (GA)-glycoside derivatives. Although two genes encoding UDP-glycosyltransferases (UGTs) that glycosylate these derivates have been functionally characterized in G. uralensis , the mechanisms of glycosylation by other UGTs remain unknown. Based on the available transcriptome data, we isolated a UGT with expression in the roots of G. uralensis . This UGT gene possibly encodes a glucosyltransferase that glycosylates GA derivatives at the 3-OH site. Biochemical analyses revealed that the recombinant UGT enzyme could transfer a glucosyl moiety to the free 3-OH or 30-COOH groups of GA. Furthermore, engineered yeast harbouring genes involved in the biosynthetic pathway for GA-glycoside derivates produced GA-3- O -β-D-glucoside, implying that the enzyme has GA 3-O-glucosyltransferase activity in vivo . Our results could provide a frame for understand the function of the UGT gene family, and also is important for further studies of triterpenoids biosynthesis in G. uralensis .

scholarly journals Genetic and Biochemical Analyses of BvgA Interaction with the Secondary Binding Region of the fhaPromoter of Bordetella pertussis

2001 ◽  
Vol 183 (2) ◽  
pp. 536-544 ◽  
Author(s):  
Philip E. Boucher ◽  
Mei-Shin Yang ◽  
Deanna M. Schmidt ◽  
Scott Stibitz
Keyword(s):  
Bordetella Pertussis ◽  
Response Regulator ◽  
Sequence Specificity ◽  
Binding Region ◽  
Dna Sequence Specificity ◽  
Genes Encoding ◽  
Two Component Signal Transduction ◽  
Biochemical Analyses

ABSTRACT The BvgA-BvgS two-component signal transduction system regulates expression of virulence factors in Bordetella pertussis. The BvgA response regulator activates transcription by binding to target promoters, which include those for the genes encoding filamentous hemagglutinin (fha) and pertussis toxin (ptx). We have previously shown that at both promoters the phosphorylated form of BvgA binds multiple high- and low-affinity sites. Specifically, at the fha promoter, we proposed that there may be high- and a low-affinity binding sites for the BvgA dimer. In our present investigation, we used DNA binding analyses and in vitro and in vivo assays of promoters with substitutions and deletions to support and extend this hypothesis. Our observations indicate that (i) binding of BvgA∼P to a primary (high-affinity) site and a secondary binding region (lower affinity) is cooperative, (ii) although both the primary binding site and the secondary binding region are required for full activity of the wild-type (undeleted) promoter, deletion of two helical turns within the secondary binding region can produce a fully active or hyperactive promoter, and (iii) BvgA binding to the secondary binding region shows limited DNA sequence specificity.

scholarly journals Transcriptomic Analysis Reveals the Mechanism of Glycyrrhizic acid Biosynthesis in Glycyrrhiza Uralensis Fisch.

2021 ◽  
Author(s):  
Junping He ◽  
Lu Yao ◽  
Juan Wang ◽  
Wenyuan Gao
Keyword(s):  
Transcription Factor ◽  
Gene Cluster ◽  
Glycyrrhizic Acid ◽  
Average Length ◽  
Glycyrrhetinic Acid ◽  
Glycyrrhiza Uralensis ◽  
Amino Acid Sequence Similarity ◽  
Acid Biosynthesis ◽  
Triterpenoid Saponins ◽  
High Amino Acid

Abstract In order to better understand the mechanism of glycyrrhizic acid biosynthesis and explore important enzyme gene resources in Glycyrrhiza uralensis Fisch., we sequenced the transcriptome of the adventitious roots of G. uralensis treated by methyl jasmonate (MJ) and assembled the de novo sequence. 256503 unique transcripts with an average length of 898bp were produced. Transcriptome sequencing and data analysis showed that the key genes of glycyrrhizic acid biosynthesis changed significantly after MJ treatment. 2720 up-regulated genes and 3493 down regulated genes were found. In the process of oxidation and glycosylation of glycyrrhizic acid biosynthesis. A putative CYP450 gene (Cluster-30944.70498) is positively correlated with glycyrrhetinic acid. The glycosyltransferase gene (Cluster-30944.25725) is positively correlated with glycyrrhizic acid and glycyrrhetinic acid. In addition, we found an AP2-EREBP family transcription factor (Cluster-30944.55070). It had high amino acid sequence similarity with PgERF1. In Panax ginseng, PgERF1 was identified as promoting the biosynthesis of triterpenoid saponins. According to the correlation analysis of transcription factors, functional gene expression and component accumulation, we speculated that this transcription factor can positively regulate the expression of farnesyl diphosphate, squalene epoxide and glycosyltransferase (Cluster-30944.25725) genes and ultimately increase the content of glycyrrhizic acid.

scholarly journals Genome-wide transcriptome analysis of triterpene biosynthetic genes of Anoectochilus roxburghii plant

2020 ◽  
Author(s):  
Hongzhen Wang ◽  
Haishun Xu ◽  
Peter E. Brodelius ◽  
Xueqian Wu ◽  
qingsong Shao ◽  
...  
Keyword(s):  
Biosynthetic Pathway ◽  
Sequence Similarity ◽  
Bioactive Components ◽  
Biosynthetic Genes ◽  
Bioactive Component ◽  
Genome Wide ◽  
Three Samples ◽  
Anoectochilus Roxburghii ◽  
Genes Encoding ◽  
Cellular Components

Abstract Background: Anoectochilus roxburghii is a medicinal plant and contains a variety of bioactive components, including triterpene, which exhibits important pharmacological properties with low toxicity. However, little is known about the biosynthetic pathway of triterpene or about the genome and transcriptome in A. roxburghii. Results: In order to analyze transcriptional determinants related to the biosynthesis of the bioactive components, we performed transcriptome sequencing in A. roxburghii (SRX1818644, SRX1818642 and SRX1818641) and annotated the sequences from three samples. In total, 137,679,059 clean reads were obtained, corresponding to 12.20 Gb of total nucleotides. They were then assembled into 86,382 contigs and 68,938 unigenes, which were further annotated according to sequence similarity with known genes in COG, EST, Nr, Pfam and Uniprot databases, leading to 10,040，29,442，39,551，34,991 and 28,082 unigenes, respectively. GO analysis classified all unigenes into three functional categories, i.e. biological processes (43,206 unigenes in 22 categories), molecular functions (46,978 unigenes in 15 categories) and cellular components (20,951 unigenes in 18 categories). Candidate triterpenes biosynthetic genes ArHMGR1 in MEV pathway, ArDXS1, ArDXS4 ArDXS5, ArDXS8-10, ArDXR1-2 and ArHDR1-2 in MEP pathway and ArFDS1, ArSM and ArOCS were selected based on RNA-seq and gene-to-metabolites correlation analysis. Conclusion: The transcriptomes of A. roxburghii plant include 86,382 contigs and 68,938 unigenes. The assembled dataset allowed identification of genes encoding enzymes in the biosynthesis of bioactive components in A. roxburghii plant. Candidate genes that encode enzymes being important in triterpenes biosynthetic pathway were selected. This will facilitate the study of expression and regulation in the biosynthesis of bioactive component in A.roxburghii.

A Comparative Pharmacokinetic Profile of Trahydropalmatine After Oral Administration of its Monomer, Rhizoma Corydalis Alkaloid Extracts and Tong-Bi-Si-Wei-Fang to Rats

2019 ◽  
Vol 15 (4) ◽  
pp. 338-345
Author(s):  
Lijun Ni ◽  
Lu Ding ◽  
Liguo Zhang ◽  
Shaorong Luan
Keyword(s):  
Oral Administration ◽  
Panax Notoginseng ◽  
Pharmacokinetic Profile ◽  
Apparent Volume ◽  
Bone Diseases ◽  
Glycyrrhiza Uralensis ◽  
Pharmacokinetic Property ◽  
Time Curve ◽  
Concentration Time

Background: Tong-Bi-Si-Wei-Fang (TBSWF) is a candidate formula of Traditional Chinese Medicine (TCM) for treating rheumatoid bone diseases, which is composed of rhizoma corydalis alkaloids, saponins of glycyrrhiza uralensis and panax notoginseng, flavonoids of rhizoma drynariae and glycyrrhiza uralensis. </P><P> Objective: Trahydropalmatine (THP), the main active ingredient of rhizoma corydalis alkaloids, was selected to study in vivo pharmacokinetics and druggability of TBSWF. Methods: The plasma concentration-time (C-T) profiles of THP and the pharmacokinetic property parameters after oral administration of THP monomer, extract of corydalis alkaloids (ECA) and TBSWF to rats, respectively were compared by a fully-validated HPLC method. Results: Compared to the THP monomer, the THP in TBSWF is absorbed faster, resides in the plasma longer and has a similar apparent volume of distribution Vz/F (10~20 L/kg). Compared to THP monomer and THP in TBSWF, the area under the concentration-time curve AUC 0-t of THP in ECA decreases two-third; Vz/F of THP in ECA (85.02 L/kg) is significantly higher than that of THP in TBSWF(p <0.05). Unlike THP monomer and THP in ECA, double peaks are observed in the C-T profile of THP after oral administration of TBSWF. THP in TBSWF exhibits slow release to a certain degree. Conclusion: The interactions among the ingredients of TBSWF promote the adsorption and prolong the residence time of THP in vivo, and provide an explanation for the advantages of TBSWF from the point of pharmacokinetics.

scholarly journals In vivo transcriptome analysis provides insights into host-dependent expression of virulence factors by Yersinia entomophaga MH96, during infection of Galleria mellonella

2020 ◽  
Vol 11 (1) ◽  
Author(s):  
Amber R Paulson ◽  
Maureen O’Callaghan ◽  
Xue-Xian Zhang ◽  
Paul B Rainey ◽  
Mark R H Hurst
Keyword(s):  
Galleria Mellonella ◽  
Functional Enrichment ◽  
Natural Environments ◽  
Insecticidal Toxin ◽  
Heat Stable ◽  
Expression Of Genes ◽  
Genes Encoding ◽  
Cell Densities

Abstract The function of microbes can be inferred from knowledge of genes specifically expressed in natural environments. Here, we report the in vivo transcriptome of the entomopathogenic bacterium Yersinia entomophaga MH96, captured during initial, septicemic, and pre-cadaveric stages of intrahemocoelic infection in Galleria mellonella. A total of 1285 genes were significantly upregulated by MH96 during infection; 829 genes responded to in vivo conditions during at least one stage of infection, 289 responded during two stages of infection, and 167 transcripts responded throughout all three stages of infection compared to in vitro conditions at equivalent cell densities. Genes upregulated during the earliest infection stage included components of the insecticidal toxin complex Yen-TC (chi1, chi2, and yenC1), genes for rearrangement hotspot element containing protein yenC3, cytolethal distending toxin cdtAB, and vegetative insecticidal toxin vip2. Genes more highly expressed throughout the infection cycle included the putative heat-stable enterotoxin yenT and three adhesins (usher-chaperone fimbria, filamentous hemagglutinin, and an AidA-like secreted adhesin). Clustering and functional enrichment of gene expression data also revealed expression of genes encoding type III and VI secretion system-associated effectors. Together these data provide insight into the pathobiology of MH96 and serve as an important resource supporting efforts to identify novel insecticidal agents.

scholarly journals Heterodimer Formation of the Homodimeric ABC Transporter OpuA

2021 ◽  
Vol 22 (11) ◽  
pp. 5912
Author(s):  
Patricia Alvarez-Sieiro ◽  
Hendrik R. Sikkema ◽  
Bert Poolman
Keyword(s):  
Abc Transporter ◽  
Conformational Dynamics ◽  
Host Organism ◽  
Affinity Tag ◽  
Practical Applications ◽  
Fundamental Research ◽  
Protein Multimerization ◽  
Biochemical Analyses

Many proteins have a multimeric structure and are composed of two or more identical subunits. While this can be advantageous for the host organism, it can be a challenge when targeting specific residues in biochemical analyses. In vitro splitting and re-dimerization to circumvent this problem is a tedious process that requires stable proteins. We present an in vivo approach to transform homodimeric proteins into apparent heterodimers, which then can be purified using two-step affinity-tag purification. This opens the door to both practical applications such as smFRET to probe the conformational dynamics of homooligomeric proteins and fundamental research into the mechanism of protein multimerization, which is largely unexplored for membrane proteins. We show that expression conditions are key for the formation of heterodimers and that the order of the differential purification and reconstitution of the protein into nanodiscs is important for a functional ABC-transporter complex.

scholarly journals Designing P. aeruginosa synthetic phages with reduced genomes

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Diana P. Pires ◽  
Rodrigo Monteiro ◽  
Dalila Mil-Homens ◽  
Arsénio Fialho ◽  
Timothy K. Lu ◽  
...  
Keyword(s):  
Galleria Mellonella ◽  
Antibacterial Properties ◽  
Genetically Engineered ◽  
In Vivo Studies ◽  
Infection Model ◽  
Promising Therapeutic Approach ◽  
Genes Encoding ◽  
First Time

AbstractIn the era where antibiotic resistance is considered one of the major worldwide concerns, bacteriophages have emerged as a promising therapeutic approach to deal with this problem. Genetically engineered bacteriophages can enable enhanced anti-bacterial functionalities, but require cloning additional genes into the phage genomes, which might be challenging due to the DNA encapsulation capacity of a phage. To tackle this issue, we designed and assembled for the first time synthetic phages with smaller genomes by knocking out up to 48% of the genes encoding hypothetical proteins from the genome of the newly isolated Pseudomonas aeruginosa phage vB_PaeP_PE3. The antibacterial efficacy of the wild-type and the synthetic phages was assessed in vitro as well as in vivo using a Galleria mellonella infection model. Overall, both in vitro and in vivo studies revealed that the knock-outs made in phage genome do not impair the antibacterial properties of the synthetic phages, indicating that this could be a good strategy to clear space from phage genomes in order to enable the introduction of other genes of interest that can potentiate the future treatment of P. aeruginosa infections.

scholarly journals The Role of Herbal Bioactive Components in Mitochondria Function and Cancer Therapy

2019 ◽  
Vol 2019 ◽  
pp. 1-12 ◽  
Author(s):  
Fangfang Tao ◽  
Yanrong Zhang ◽  
Zhiqian Zhang
Keyword(s):  
Cancer Therapy ◽  
Cancer Cell ◽  
Apoptotic Cell ◽  
Redox Status ◽  
Cancer Therapies ◽  
Bioactive Components ◽  
Atp Production

Mitochondria are highly dynamic double-membrane organelles which play a well-recognized role in ATP production, calcium homeostasis, oxidation-reduction (redox) status, apoptotic cell death, and inflammation. Dysfunction of mitochondria has long been observed in a number of human diseases, including cancer. Targeting mitochondria metabolism in tumors as a cancer therapeutic strategy has attracted much attention for researchers in recent years due to the essential role of mitochondria in cancer cell growth, apoptosis, and progression. On the other hand, a series of studies have indicated that traditional medicinal herbs, including traditional Chinese medicines (TCM), exert their potential anticancer effects as an effective adjunct treatment for alleviating the systemic side effects of conventional cancer therapies, for reducing the risk of recurrence and cancer mortality and for improving the quality of patients’ life. An amazing feature of these structurally diverse bioactive components is that majority of them target mitochondria to provoke cancer cell-specific death program. The aim of this review is to summarize the in vitro and in vivo studies about the role of these herbs, especially their bioactive compounds in the modulation of the disturbed mitochondrial function for cancer therapy.

18β-Glycyrrhetinic Acid, a Novel Naturally Derived Agent, Suppresses Prolactin Hyperactivity and Reduces Antipsychotic-Induced Hyperprolactinemia in In Vitro and In Vivo Models

2016 ◽  
Vol 41 (9) ◽  
pp. 2233-2242 ◽  
Author(s):  
Di Wang ◽  
Yongfeng Zhang ◽  
Chunyue Wang ◽  
Dongxu Jia ◽  
Guangsheng Cai ◽  
...  
Keyword(s):  
Glycyrrhetinic Acid ◽  
In Vivo Models
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