scholarly journals Identification of a Novel Specific Cucurbitadienol Synthase Allele in Siraitia grosvenorii Correlates with High Catalytic Efficiency

Molecules ◽  
2019 ◽  
Vol 24 (3) ◽  
pp. 627 ◽  
Author(s):  
Jing Qiao ◽  
Zuliang Luo ◽  
Zhe Gu ◽  
Yanling Zhang ◽  
Xindan Zhang ◽  
...  
Keyword(s):  
Molecular Breeding ◽  
Specific Activity ◽  
Biological Activities ◽  
Enzyme Reaction ◽  
Catalytic Efficiency ◽  
Wild Type ◽  
Siraitia Grosvenorii ◽  
Wide Range ◽  
Triterpenoid Glycosides

Mogrosides, the main bioactive compounds isolated from the fruits of Siraitia grosvenorii, are a group of cucurbitane-type triterpenoid glycosides that exhibit a wide range of notable biological activities and are commercially available worldwide as natural sweeteners. However, the extraction cost is high due to their relatively low contents in plants. Therefore, molecular breeding needs to be achieved when conventional plant breeding can hardly improve the quality so far. In this study, the levels of 21 active mogrosides and two precursors in 15 S. grosvenorii varieties were determined by HPLC-MS/MS and GC-MS, respectively. The results showed that the variations in mogroside V content may be caused by the accumulation of cucurbitadienol. Furthermore, a total of four wild-type cucurbitadienol synthase protein variants (50R573L, 50C573L, 50R573Q, and 50C573Q) based on two missense mutation single nucleotide polymorphism (SNP) sites were discovered. An in vitro enzyme reaction analysis indicated that 50R573L had the highest activity, with a specific activity of 10.24 nmol min−1 mg−1. In addition, a site-directed mutant, namely, 50K573L, showed a 33% enhancement of catalytic efficiency compared to wild-type 50R573L. Our findings identify a novel cucurbitadienol synthase allele correlates with high catalytic efficiency. These results are valuable for the molecular breeding of luohanguo.

scholarly journals Improving the Stability and Activity of Arginine Decarboxylase at Alkaline pH for the Production of Agmatine

2021 ◽  
Vol 1 ◽  
Author(s):  
Eun Young Hong ◽  
Sun-Gu Lee ◽  
Hyungdon Yun ◽  
Byung-Gee Kim
Keyword(s):  
Disulfide Bond ◽  
Specific Activity ◽  
Catalytic Efficiency ◽  
Arginine Decarboxylase ◽  
Saturation Mutagenesis ◽  
Alkaline Ph ◽  
Wild Type ◽  
Active Site Residues ◽  
Cellular Mechanisms

Agmatine, involved in various modulatory actions in cellular mechanisms, is produced from arginine (Arg) by decarboxylation reaction using arginine decarboxylase (ADC, EC 4.1.1.19). The major obstacle of using wild-type Escherichia coli ADC (ADCes) in agmatine production is its sharp activity loss and instability at alkaline pH. Here, to overcome this problem, a new disulfide bond was rationally introduced in the decameric interface region of the enzyme. Among the mutants generated, W16C/D43C increased both thermostability and activity. The half-life (T1/2) of W16C/D43C at pH 8.0 and 60°C was 560 min, which was 280-fold longer than that of the wild-type, and the specific activity at pH 8.0 also increased 2.1-fold. Site-saturation mutagenesis was subsequently performed at the active site residues of ADCes using the disulfide-bond mutant (W16C/D43C) as a template. The best variant W16C/D43C/I258A displayed a 4.4-fold increase in the catalytic efficiency when compared with the wild-type. The final mutant (W16C/D43C/I258A) was successfully applied to in vitro synthesis of agmatine with an improved yield and productivity (>89.0% yield based on 100 mM of Arg within 5  h).

Chalcones: As potent α-amylase enzyme inhibitors; synthesis, in vitro, and in silico studies

2020 ◽  
Vol 16 ◽  
Author(s):  
Mahboob Ali ◽  
Momin Khan ◽  
Khair Zaman ◽  
Abdul Wadood ◽  
Maryam Iqbal ◽  
...  
Keyword(s):  
In Silico ◽  
Enzyme Inhibitors ◽  
Biological Activities ◽  
Condensation Reaction ◽  
Type Ii Diabetes Mellitus ◽  
Spectroscopic Techniques ◽  
Chalcone Derivatives ◽  
In Silico Studies ◽  
Wide Range

: Background: The inhibition of α-amylase enzyme is one of the best therapeutic approach for the management of type II diabetes mellitus. Chalcone possesses a wide range of biological activities. Objective: In the current study chalcone derivatives (1-17) were synthesized and evaluated their inhibitory potential against α-amylase enzyme. Method: For that purpose, a library of substituted (E)-1-(naphthalene-2-yl)-3-phenylprop-2-en-1-ones was synthesized by ClaisenSchmidt condensation reaction of 2-acetonaphthanone and substituted aryl benzaldehyde in the presence of base and characterized via different spectroscopic techniques such as EI-MS, HREI-MS, 1H-, and 13C-NMR. Results: Sixteen synthetic chalcones were evaluated for in vitro porcine pancreatic α-amylase inhibition. All the chalcones demonstrated good inhibitory activities in the range of IC50 = 1.25 ± 1.05 to 2.40 ± 0.09 μM as compared to the standard commercial drug acarbose (IC50 = 1.34 ± 0.3 μM). Conclusion: Chalcone derivatives (1-17) were synthesized, characterized, and evaluated for their α-amylase inhibition. SAR revealed that electron donating groups in the phenyl ring have more influence on enzyme inhibition. However, to insight the participation of different substituents in the chalcones on the binding interactions with the α-amylase enzyme, in silico (computer simulation) molecular modeling analyses were carried out.

Associated microflora of medicinal ferns: biotechnological potentials and possible applications

2016 ◽  
Vol 5 (03) ◽  
pp. 4927 ◽  
Author(s):  
Shubhi Srivastava ◽  
Paul A. K.
Keyword(s):  
Secondary Metabolites ◽  
Growth Promotion ◽  
Biological Activities ◽  
Hydrolytic Enzymes ◽  
In Vitro Production ◽  
Wide Range ◽  
Negative Effect ◽  
Bacteria And Fungi

Plant associated microorganisms that colonize the upper and internal tissues of roots, stems, leaves and flowers of healthy plants without causing any visible harmful or negative effect on their host. Diversity of microbes have been extensively studied in a wide variety of vascular plants and shown to promote plant establishment, growth and development and impart resistance against pathogenic infections. Ferns and their associated microbes have also attracted the attention of the scientific communities as sources of novel bioactive secondary metabolites. The ferns and fern alleles, which are well adapted to diverse environmental conditions, produce various secondary metabolites such as flavonoids, steroids, alkaloids, phenols, triterpenoid compounds, variety of amino acids and fatty acids along with some unique metabolites as adaptive features and are traditionally used for human health and medicine. In this review attention has been focused to prepare a comprehensive account of ethnomedicinal properties of some common ferns and fern alleles. Association of bacteria and fungi in the rhizosphere, phyllosphere and endosphere of these medicinally important ferns and their interaction with the host plant has been emphasized keeping in view their possible biotechnological potentials and applications. The processes of host-microbe interaction leading to establishment and colonization of endophytes are less-well characterized in comparison to rhizospheric and phyllospheric microflora. However, the endophytes are possessing same characteristics as rhizospheric and phyllospheric to stimulate the in vivo synthesis as well as in vitro production of secondary metabolites with a wide range of biological activities such as plant growth promotion by production of phytohormones, siderophores, fixation of nitrogen, and phosphate solubilization. Synthesis of pharmaceutically important products such as anticancer compounds, antioxidants, antimicrobials, antiviral substances and hydrolytic enzymes could be some of the promising areas of research and commercial exploitation.

Combinatorial Synthesis of Novel 1-Sulfonyloxy/acyloxyeugenol Derivatives as Fungicidal Agents

2021 ◽  
Vol 24 ◽  
Author(s):  
Genqiang Chen ◽  
Lina Zhu ◽  
Jiaxuan He ◽  
Song Zhang ◽  
Yuanhao Li ◽  
...  
Keyword(s):  
Fungicidal Activity ◽  
High Efficiency ◽  
Biological Activities ◽  
Phytophthora Capsici ◽  
Effective Control ◽  
Syzygium Aromaticum ◽  
Wide Range ◽  
Structural Skeleton ◽  
Acyloxy Group

Background: Developing the high-efficiency and low-risk small-molecule green-fungicide is the key to effective control of the plant pathogenic oomycetes. Essential oils play a very important role in novel fungicide discovery for their unique sources and potential target sites. Eugenol, a kind of plant essential oil, was mainly isolated from the unopened and dried flower buds of Syzygium aromaticum of the Myrtaceae family. Due to its unique structural skeleton, eugenol and its derivatives have exhibited a wide range of biological activities. However, study on the synthesis of novel 1-sulfonyloxy/acyloxyeugenol derivatives as fungicidal agents against Phytophthora capsici has not yet been reported. Methods: Twenty-six novel 1-sulfonyloxy/acyloxyeugenol derivatives (3a-p and 5a-j) were prepared and their structures were well characterized by 1H NMR, HRMS, and m.p.. Their fungicidal activity was evaluated against P. capsici by using the mycelial growth rate method. Results: To find novel natural-product-based fungicidal agents to control the plant pathogenic oomycetes, we herein designed and synthesized two series of novel 1-sulfonyloxy/acyloxyeugenol derivatives (3a-p and 5a-j) as fungicidal agents against P. capsici Leonian, in vitro. Results of fungicidal activity revealed that, among all compounds, especially compounds 3a, 3f, and 3n displayed the most potent anti-oomycete activity against P. capsici with EC50 values of 79.05, 75.05, and 70.80, respectively. Conclusion: The results revealed that the anti-oomycete activity of eugenol with the sulfonyloxy group was higher than that with the acyloxy group. It is suggested that the fungicidal activity of eugenol can be improved by introducing the sulfonyloxy group. This will pave the way for further design, structural modification, and to develop eugenol derivatives as fungicidal agents.

scholarly journals In vitro antimicrobial activity of the organic extract of Cladonia substellata Vainio and usnic acid against Staphylococcus spp. obtained from cats and dogs

2017 ◽  
Vol 37 (4) ◽  
pp. 368-378 ◽  
Author(s):  
Jusciêne B. Moura ◽  
Agueda C. de Vargas ◽  
Gisele V. Gouveia ◽  
João J. de S. Gouveia ◽  
Juracy C. Ramos-Júnior ◽  
...  
Keyword(s):  
Antimicrobial Activity ◽  
Biological Activities ◽  
Meca Gene ◽  
Usnic Acid ◽  
Organic Extract ◽  
In Vitro Antimicrobial Activity ◽  
Low Concentrations ◽  
Wide Range ◽  
Staphylococcus Spp

ABSTRACT: Cladonia substellata Vainio is a lichen found in different regions of the world, including the Northeast of Brazil. It contains several secondary metabolites with biological activity, including usnic acid, which has exhibited a wide range of biological activities. The aim of this study was to evaluate the in vitro antimicrobial activity of the organic extract of C. substellata and purified usnic acid. Initially, Staphylococcus spp., derived from samples of skin and ears of dogs and cats with suspected pyoderma and otitis, were isolated and analyzed. In antimicrobial susceptibility testing against Staphylococcus spp., 77% (105/136) of the isolates were resistant to the antimicrobials tested. In the assessment of biofilm production, 83% (113/136) were classified as producing biofilm. In genetic characterization, 32% (44/136) were positive for blaZ, no isolate (0/136) was positive for the mecA gene, and 2% (3/136) were positive for the icaD gene. The in vitro antimicrobial activity of the organic extract of C. substellata and purified usnic acid against Staphylococcus spp. ranged from 0.25mg/mL to 0.0019mg/mL, inhibiting bacterial growth at low concentrations. The substances were more effective against biofilm-producing bacteria (0.65mg/mL-0.42mg/mL) when compared to non-biofilm producing bacteria (2.52mg/mL-2.71mg/mL). Usnic acid and the organic extract of C. substellata can be effective in the treatment of pyoderma and otitis in dogs and cats caused by Staphylococcus spp.

scholarly journals Genome-Wide fitness analysis of group B Streptococcus in human amniotic fluid reveals a transcription factor that controls multiple virulence traits

2021 ◽  
Vol 17 (3) ◽  
pp. e1009116
Author(s):  
Allison N. Dammann ◽  
Anna B. Chamby ◽  
Andrew J. Catomeris ◽  
Kyle M. Davidson ◽  
Hervé Tettelin ◽  
...  
Keyword(s):  
Transcription Factor ◽  
Amniotic Fluid ◽  
Deletion Strain ◽  
Growth Defect ◽  
Wild Type ◽  
Human Amniotic Fluid ◽  
Wide Range ◽  
Pregnant Mice ◽  
Group B

Streptococcus agalactiae (group B Streptococcus; GBS) remains a dominant cause of serious neonatal infections. One aspect of GBS that renders it particularly virulent during the perinatal period is its ability to invade the chorioamniotic membranes and persist in amniotic fluid, which is nutritionally deplete and rich in fetal immunologic factors such as antimicrobial peptides. We used next-generation sequencing of transposon-genome junctions (Tn-seq) to identify five GBS genes that promote survival in the presence of human amniotic fluid. We confirmed our Tn-seq findings using a novel CRISPR inhibition (CRISPRi) gene expression knockdown system. This analysis showed that one gene, which encodes a GntR-class transcription factor that we named MrvR, conferred a significant fitness benefit to GBS in amniotic fluid. We generated an isogenic targeted deletion of the mrvR gene, which had a growth defect in amniotic fluid relative to the wild type parent strain. The mrvR deletion strain also showed a significant biofilm defect in vitro. Subsequent in vivo studies showed that while the mutant was able to cause persistent murine vaginal colonization, pregnant mice colonized with the mrvR deletion strain did not develop preterm labor despite consistent GBS invasion of the uterus and the fetoplacental units. In contrast, pregnant mice colonized with wild type GBS consistently deliver prematurely. In a sepsis model the mrvR deletion strain showed significantly decreased lethality. In order to better understand the mechanism by which this newly identified transcription factor controls GBS virulence, we performed RNA-seq on wild type and mrvR deletion GBS strains, which revealed that the transcription factor affects expression of a wide range of genes across the GBS chromosome. Nucleotide biosynthesis and salvage pathways were highly represented among the set of differentially expressed genes, suggesting that MrvR may be involved in regulating nucleotide availability.

scholarly journals Mutational Analysis of Ocriplasmin to Reduce Proteolytic and Autolytic Activity in Pichia pastoris

2020 ◽  
Vol 22 (1) ◽  
Author(s):  
Roghayyeh Baghban ◽  
Safar Farajnia ◽  
Younes Ghasemi ◽  
Reyhaneh Hoseinpoor ◽  
Azam Safary ◽  
...  
Keyword(s):  
Pichia Pastoris ◽  
Proteolytic Activity ◽  
Mutational Analysis ◽  
Expression System ◽  
Catalytic Efficiency ◽  
Site Directed Mutagenesis ◽  
Wild Type ◽  
Vitreomacular Adhesion ◽  
Autolytic Activity

Abstract Background Ocriplasmin (Jetrea) is using for the treatment of symptomatic vitreomacular adhesion. This enzyme undergoes rapid inactivation and limited activity duration as a result of its autolytic nature after injection within the eye. Moreover, the proteolytic activity can cause photoreceptor damage, which may result in visual impairment in more serious cases. Results The present research aimed to reduce the disadvantages of ocriplasmin using site-directed mutagenesis. To reduce the autolytic activity of ocriplasmin in the first variant, lysine 156 changed to glutamic acid and, in the second variant for the proteolytic activity reduction, alanine 59 mutated to threonine. The third variant contained both mutations. Expression of wild type and three mutant variants of ocriplasmin constructs were done in the Pichia pastoris expression system. The mutant variants were analyzed in silico and in vitro and compared to the wild type. The kinetic parameters of ocriplasmin variants showed both variants with K156E substitution were more resistant to autolytic degradation than wild-type. These variants also exhibited reduced Kcat and Vmax values. An increase in their Km values, leading to a decreased catalytic efficiency (the Kcat/Km ratio) of autolytic and mixed variants. Moreover, in the variant with A59T mutation, Kcat and Vmax values have reduced compared to wild type. The mix variants showed the most increase in Km value (almost 2-fold) as well as reduced enzymatic affinity to the substrate. Thus, the results indicated that combined mutations at the ocriplasmin sequence were more effective compared with single mutations. Conclusions The results indicated such variants represent valuable tools for the investigation of therapeutic strategies aiming at the non-surgical resolution of vitreomacular adhesion.

scholarly journals Enhanced Recovery of Antioxidant Compounds from Hazelnut (Corylus avellana L.) Involucre Based on Extraction Optimization: Phytochemical Profile and Biological Activities

Antioxidants ◽  
2019 ◽  
Vol 8 (10) ◽  
pp. 460 ◽  
Author(s):  
Rusu ◽  
Fizeșan ◽  
Pop ◽  
Gheldiu ◽  
Mocan ◽  
...  
Keyword(s):  
Bioactive Compounds ◽  
Enhanced Recovery ◽  
Biological Activities ◽  
Natural Antioxidants ◽  
Antioxidant Compounds ◽  
Cancerous Cell ◽  
Wide Range ◽  
Quantitative Analyses ◽  
Corylus Avellana L

Tree nut by-products could contain a wide range of phytochemicals, natural antioxidants, which might be used as a natural source for dietary supplements. The aim of the present study was to evaluate the phenolic and sterolic composition, as well as the antioxidant and other biological activities, of hazelnut involucre (HI) extracts. Experimental designs were developed in order to select the optimum extraction conditions (solvent, temperature, time) using turbo-extraction by Ultra-Turrax for obtaining extracts rich in bioactive compounds. Qualitative and quantitative analyses were performed by LC-MS and LC-MS/MS and they revealed important amounts of individual polyphenols and phytosterols, molecules with antioxidant potential. The richest polyphenolic HI extract with the highest antioxidant activity by TEAC assay was further evaluated by other in vitro antioxidant tests (DPPH, FRAP) and enzyme inhibitory assays. Additionally, the cytotoxic and antioxidant effects of this extract on two cancerous cell lines and on normal cells were tested. This is the first study to analyze the composition of both hydrophilic and lipophilic bioactive compounds in HI extracts. Our findings reveal that this plant by-product presents strong biological activities, justifying further research, and it could be considered an inexpensive source of natural antioxidants for food, pharmaceutical, or cosmetic industry.

scholarly journals Inclusion Complexes of β and HPβ-Cyclodextrin with α, β Amyrin and In Vitro Anti-Inflammatory Activity

Biomolecules ◽  
2019 ◽  
Vol 9 (6) ◽  
pp. 241 ◽  
Author(s):  
Walter Ferreira da Silva Júnior ◽  
Danielle Lima Bezerra de Menezes ◽  
Luana Carvalho de Oliveira ◽  
Letícia Scherer Koester ◽  
Patrícia Danielle Oliveira de Almeida ◽  
...  
Keyword(s):  
Inclusion Complexes ◽  
Biological Activities ◽  
Physicochemical Characterization ◽  
Inflammatory Activity ◽  
Scanning Calorimetry ◽  
Promising Alternative ◽  
Wide Range ◽  
Natural Mixture ◽  
Anti Inflammatory

α, β amyrin (ABAM) is a natural mixture of pentacyclic triterpenes that has a wide range of biological activities. ABAM is isolated from the species of the Burseraceae family, in which the species Protium is commonly found in the Amazon region of Brazil. The aim of this work was to develop inclusion complexes (ICs) of ABAM and β-cyclodextrin (βCD) and hydroxypropyl-β-cyclodextrin (HPβCD) by physical mixing (PM) and kneading (KN) methods. Interactions between ABAM and the CD’s as well as the formation of ICs were confirmed by physicochemical characterization in the solid state by Fourier transform infrared (FTIR), scanning electron microscopy (SEM), X-ray diffraction (XRD), thermogravimetry (TG) and differential scanning calorimetry (DSC). Physicochemical characterization indicated the formation of ICs with both βCD and HPβCD. Such ICs were able to induce changes in the physicochemical properties of ABAM. In addition, the formation of ICs with cyclodextrins showed to be an effective and promising alternative to enhance the anti-inflammatory activity and safety of ABAM.

scholarly journals Targeting mutant p53-expressing tumours with a T cell receptor-like antibody specific for a wild-type antigen

2019 ◽  
Vol 10 (1) ◽  
Author(s):  
Lionel Low ◽  
Angeline Goh ◽  
Joanna Koh ◽  
Samantha Lim ◽  
Cheng-I Wang
Keyword(s):  
Cell Receptor ◽  
Class I ◽  
Human Antibody ◽  
Mutant P53 ◽  
Wild Type ◽  
Class I Mhc ◽  
Type Antigen ◽  
Wide Range

AbstractAccumulation of mutant p53 proteins is frequently found in a wide range of cancers. While conventional antibodies fail to target intracellular proteins, proteosomal degradation results in the presentation of p53-derived peptides on the tumour cell surface by class I molecules of the major histocompatibility complex (MHC). Elevated levels of such p53-derived peptide-MHCs on tumour cells potentially differentiate them from healthy tissues. Here, we report the engineering of an affinity-matured human antibody, P1C1TM, specific for the unmutated p53125-134 peptide in complex with the HLA-A24 class I MHC molecule. We show that P1C1TM distinguishes between mutant and wild-type p53 expressing HLA-A24+ cells, and mediates antibody dependent cellular cytotoxicity of mutant p53 expressing cells in vitro. Furthermore, we show that cytotoxic PNU-159682-P1C1TM drug conjugates specifically inhibit growth of mutant p53 expressing cells in vitro and in vivo. Hence, p53-associated peptide-MHCs are attractive targets for the immunotherapy against mutant p53 expressing tumours.

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