scholarly journals Peptides Derived From the α-Core and γ-Core Regions of a Putative Silybum marianum Flower Defensin Show Antifungal Activity Against Fusarium graminearum

2021 ◽  
Vol 12 ◽  
Author(s):  
Agustina Fernández ◽  
María Laura Colombo ◽  
Lucrecia M. Curto ◽  
Gabriela E. Gómez ◽  
José M. Delfino ◽  
...  
Keyword(s):  
Fusarium Graminearum ◽  
Core Region ◽  
Terminal Region ◽  
Wild Plant ◽  
Wheat Crop ◽  
Antimicrobial Protein ◽  
Ionization Mass ◽  
Silybum Marianum ◽  
Plant Defensins ◽  
Antifungal Peptides

Fusarium graminearum is the etiological agent of Fusarium head blight (FHB), a disease that produces a significant decrease in wheat crop yield and it is further aggravated by the presence of mycotoxins in the affected grains that may cause health problems to humans and animals. Plant defensins and defensin-like proteins are antimicrobial peptides (AMPs); they are small basic, cysteine-rich peptides (CRPs) ubiquitously expressed in the plant kingdom and mostly involved in host defence. They present a highly variable sequence but a conserved structure. The γ-core located in the C-terminal region of plant defensins has a conserved β-hairpin structure and is a well-known determinant of the antimicrobial activity among disulphide-containing AMPs. Another conserved motif of plant defensins is the α-core located in the N-terminal region, not conserved among the disulphide-containing AMPs, it has not been yet extensively studied. In this report, we have cloned the putative antimicrobial protein DefSm2, expressed in flowers of the wild plant Silybum marianum. The cDNA encodes a protein with two fused basic domains of an N-terminal defensin domain (DefSm2-D) and a C-terminal Arg-rich and Lys-rich domain. To further characterize the DefSm2-D domain, we built a 3D template-based model that will serve to support the design of novel antifungal peptides. We have designed four potential antifungal peptides: two from the DefSm2-D α-core region (SmAPα1-21 and SmAPα10-21) and two from the γ-core region (SmAPγ27-44 and SmAPγ29-35). We have chemically synthesized and purified the peptides and further characterized them by electrospray ionization mass spectrometry (ESI-MS) and Circular dichroism (CD) spectroscopy. SmAPα1-21, SmAPα10-21, and SmAPγ27-44 inhibited the growth of the phytopathogen F. graminearum at low micromolar concentrations. Conidia exposure to the fungicidal concentration of the peptides caused membrane permeabilization to the fluorescent probe propidium iodide (PI), suggesting that this is one of the main contributing factors in fungal cell killing. Furthermore, conidia treated for 0.5h showed cytoplasmic disorganization as observed by transmission electron microscopy (TEM). Remarkably, the peptides derived from the α-core induced morphological changes on the conidia cell wall, which is a promising target since its distinctive biochemical and structural organization is absent in plant and mammalian cells.

scholarly journals TOTAL POLYPHENOLIC CONTENT AND ANTIOXIDANT ACTIVITY OF SILYBUM MARIANUM CULTURES GROWN ON DIFFERENT GROWTH REGULATORS

2017 ◽  
Vol 9 (4) ◽  
pp. 44 ◽  
Author(s):  
Hattem Mekky ◽  
Samah El Sohafy ◽  
Rasha Abu El-khair ◽  
Amr El Hawiet
Keyword(s):  
Antioxidant Activity ◽  
Gallic Acid ◽  
Growth Regulators ◽  
Phenolic Content ◽  
Wild Plant ◽  
Total Phenolic ◽  
Silybum Marianum ◽  
Polyphenolic Content ◽  
Trolox Equivalent ◽  
Total Polyphenolic Content

Objective: The present study was designed to study the effect of supplementing Silybum marianum cultures with different combinations and concentrations of growth regulators on the total phenolic content and the antioxidant activity of the produced cultures.Methods: The total polyphenolic content of 23 extracts of calli was calculated as µg of gallic acid/mg of dry weight extract (DWE) using Folin–Ciocalteu reagent. The antioxidant activity was calculated as trolox equivalent antioxidant capacity using 1,1-Diphenyl-2-picryl-hydrazyl (DPPH) assay for extracts containing more than 10 µg gallic acid/mg DWE.Results: Most of the combinations showed an increase in the total polyphenolic concentration compared to the wild plant. However, cultures grown on media supplemented with combinations of benzyl aminopurine (BAP) and gebberellic acid (Gb), adenine (Ad) and kinetin (Kin), BAP and indole acetic acid (IAA) possessed the highest total polyphenolic contents 20.5, 13.09 and 12.15 µg gallic acid/mg DWE, respectively. However, only (BAP+Gb), (BAP+1-Naphthaleneacetic acid (NAA), IAA, kin and (BAP+Ad) showed a significant increase in the antioxidant activity calculated as trolox equivalent with 2.65, 1.56, 1.56, 1.35 and 1.22 folds increase over the wild plant, respectively.Conclusion: The results obtained clearly indicated that changing the growth regulator system of Silybum marianum cultures serves as an easy and reliable method for increasing the total phenolic content and the antioxidant activity of cultures.

scholarly journals Sphingolipid C-9 Methyltransferases Are Important for Growth and Virulence but Not for Sensitivity to Antifungal Plant Defensins in Fusarium graminearum

2008 ◽  
Vol 8 (2) ◽  
pp. 217-229 ◽  
Author(s):  
Vellaisamy Ramamoorthy ◽  
Edgar B. Cahoon ◽  
Mercy Thokala ◽  
Jagdeep Kaur ◽  
Jia Li ◽  
...  
Keyword(s):  
Fusarium Graminearum ◽  
Type Strain ◽  
Wild Type Strain ◽  
Fungal Growth ◽  
Wild Type ◽  
Knockout Mutant ◽  
Double Knockout ◽  
Disease Symptoms ◽  
Plant Defensins ◽  
Antifungal Action

ABSTRACT The C-9-methylated glucosylceramides (GlcCers) are sphingolipids unique to fungi. They play important roles in fungal growth and pathogenesis, and they act as receptors for some antifungal plant defensins. We have identified two genes, FgMT1 and FgMT2, that each encode a putative sphingolipid C-9 methyltransferase (C-9-MT) in the fungal pathogen Fusarium graminearum and complement a Pichia pastoris C-9-MT-null mutant. The ΔFgmt1 mutant produced C-9-methylated GlcCer like the wild-type strain, PH-1, whereas the ΔFgmt2 mutant produced 65 to 75% nonmethylated and 25 to 35% methylated GlcCer. No ΔFgmt1ΔFgmt2 double-knockout mutant producing only nonmethylated GlcCer could be recovered, suggesting that perhaps C-9-MTs are essential in this pathogen. This is in contrast to the nonessential nature of this enzyme in the unicellular fungus P. pastoris. The ΔFgmt2 mutant exhibited severe growth defects and produced abnormal conidia, while the ΔFgmt1 mutant grew like the wild-type strain, PH-1, under the conditions tested. The ΔFgmt2 mutant also exhibited drastically reduced disease symptoms in wheat and much-delayed disease symptoms in Arabidopsis thaliana. Surprisingly, the ΔFgmt2 mutant was less virulent on different host plants tested than the previously characterized ΔFggcs1 mutant, which lacks GlcCer synthase activity and produces no GlcCer at all. Moreover, the ΔFgmt1 and ΔFgmt2 mutants, as well as the P. pastoris strain in which the C-9-MT gene was deleted, retained sensitivity to the antifungal plant defensins MsDef1 and RsAFP2, indicating that the C-9 methyl group is not a critical structural feature of the GlcCer receptor required for the antifungal action of plant defensins.

scholarly journals Green Bio-Assisted Synthesis, Characterization and Biological Evaluation of Biocompatible ZnO NPs Synthesized from Different Tissues of Milk Thistle (Silybum marianum)

Nanomaterials ◽  
2019 ◽  
Vol 9 (8) ◽  
pp. 1171 ◽  
Author(s):  
Bilal Haider Abbasi ◽  
Muzamil Shah ◽  
Syed Salman Hashmi ◽  
Munazza Nazir ◽  
Sania Naz ◽  
...  
Keyword(s):  
Thermo Gravimetric Analysis ◽  
Radical Scavenging ◽  
Biological Evaluation ◽  
Seed Extract ◽  
Wild Plant ◽  
Gravimetric Analysis ◽  
Silybum Marianum ◽  
Zno Nps ◽  
Wide Range ◽  
Different Tissues

The purpose of the current study was green synthesis of ZnO-nanoparticles (NPs) from different tissues of Silybum marianum (L.) Gaernt. (i.e., seeds, wild plant, in vitro derived plantlets and callus cultures) followed by extensive characterization and evaluation of their biological potency. ZnO-NPs thus synthesized were subjected to characterization using standard techniques such as XRD, FTIR and SEM. Thermal stability of synthesized NPs was also evaluated using thermo-gravimetric analysis. Highly stable crystalline NPs with size ranging between 30.8 and 46.0 nm were obtained from different tissues of S. marianum. These NPs have revealed a wide range of biological applications showing antioxidant, moderate α-amylase inhibitor, antibacterial and cytotoxic potencies. The highest antibacterial activity (20 ± 0.98 mm) was shown by seed extract-mediated ZnO NPs against Staphylococcus aureus (ATCC-6538). Seed extract-mediated ZnO NPs also showed the most potent antioxidant activity (27.7 ± 0.9 µgAAE/mg, 23.8 ± 0.7 µgAAE/mg and 12.7 ± 1.9% total antioxidant capacity (TAC), total reducing power (TRP) and DPPH-free radical scavenging assay (FRSA), respectively). All of the synthesized ZnO NPs also showed cytotoxic activity against the hepato-cellular carcinoma (HepG2) human cells. Interestingly, these ZnO NPs were also highly biocompatible, as evidenced by the brine shrimp lethality and human red blood cells hemolytic assays. Among all of the NPs synthesized and used, the effect of seed extract-mediated NPs was found to be most promising for future applications.

scholarly journals Transducin Beta-Like Gene FTL1 Is Essential for Pathogenesis in Fusarium graminearum

2009 ◽  
Vol 8 (6) ◽  
pp. 867-876 ◽  
Author(s):  
Shengli Ding ◽  
Rahim Mehrabi ◽  
Cornelia Koten ◽  
Zhensheng Kang ◽  
Yangdou Wei ◽  
...  
Keyword(s):  
Fusarium Graminearum ◽  
Insertional Mutagenesis ◽  
Critical Role ◽  
Histone Deacetylation ◽  
Head Blight ◽  
Plant Infection ◽  
Expression Of Genes ◽  
Plant Defensins ◽  
Mitogen Activated Protein ◽  
Late Stages

ABSTRACT Fusarium head blight caused by Fusarium graminearum is an important disease of wheat and barley. In a previous study, we identified several mutants with reduced virulence by insertional mutagenesis. A transducin beta-like gene named FTL1 was disrupted in one of these nonpathogenic mutants. FTL1 is homologous to Saccharomyces cerevisiae SIF2, which is a component of the Set3 complex involved in late stages of ascospore formation. The Δftl1 mutant was significantly reduced in conidiation and failed to cause typical disease symptoms. It failed to colonize the vascular tissues of rachis or cause necrosis on the rachis of inoculated wheat heads. The Δftl1 mutant also was defective in spreading from infected anthers to ovaries and more sensitive than the wild type to plant defensins MsDef1 and osmotin. However, the activation of two mitogen-activated protein kinases, Mgv1 and Gpmk1, production of deoxynivalenol, and expression of genes known to be important for plant infection in F. graminearum were not affected, indicating that the defect of the Δftl1 mutant in plant infection is unrelated to known virulence factors in this pathogen and may involve novel mechanisms. The Δftl1 deletion mutant was significantly reduced in histone deacetylation, and many members of the yeast Set3 complex are conserved in F. graminearum. FTL1 appears to be a component of this well-conserved protein complex that plays a critical role in the penetration and colonization of wheat tissues.

scholarly journals Bioconversion of Callus-Produced Precursors to Silymarin Derivatives in Silybum marianum Leaves for the Production of Bioactive Compounds

2021 ◽  
Vol 22 (4) ◽  
pp. 2149
Author(s):  
Dina Gad ◽  
Hamed El-Shora ◽  
Daniele Fraternale ◽  
Elisa Maricchiolo ◽  
Andrea Pompa ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Mass Spectra ◽  
Phenylpropanoid Pathway ◽  
High Yield ◽  
Ionization Mass ◽  
Silybum Marianum ◽  
Metabolic Capacity ◽  
Chemopreventive Agent ◽  
Caffeoylquinic Acid ◽  
Full Capacity

The present study aimed to investigate the enzymatic potential of Silybum marianum leaves to bioconvert phenolic acids produced in S. marianum callus into silymarin derivatives as chemopreventive agent. Here we demonstrate that despite the fact that leaves of S. marianum did not accumulate silymarin themselves, expanding leaves had the full capacity to convert di-caffeoylquinic acid to silymarin complex. This was proven by HPLC separations coupled with electrospray ionization mass spectrometry (ESI-MS) analysis. Soaking the leaf discs with S. marianum callus extract for different times revealed that silymarin derivatives had been formed at high yield after 16 h. Bioconverted products displayed the same retention time and the same mass spectra (MS or MS/MS) as standard silymarin. Bioconversion was achieved only when using leaves of a specific age, as both very young and old leaves failed to produce silymarin from callus extract. Only medium leaves had the metabolic capacity to convert callus components into silymarin. The results revealed higher activities of enzymes of the phenylpropanoid pathway in medium leaves than in young and old leaves. It is concluded that cotyledon-derived callus efficiently produces compounds that can be bio-converted to flavonolignans in leaves tissue of S. marianum.

scholarly journals Interaction between N-terminal region and β core region in photoactive yellow protein.

2003 ◽  
Vol 43 (supplement) ◽  
pp. S196
Author(s):  
H. Kamikubo ◽  
N. Shimizu ◽  
M. Harigai ◽  
Y. Yamazaki ◽  
Y. Imamoto ◽  
...  
Keyword(s):  
Core Region ◽  
Terminal Region ◽  
Photoactive Yellow Protein

scholarly journals Acylation-stimulating protein (ASP): structure–function determinants of cell surface binding and triacylglycerol synthetic activity

1999 ◽  
Vol 342 (1) ◽  
pp. 41-48 ◽  
Author(s):  
Ian MURRAY ◽  
Jörg KÖHL ◽  
Katherine CIANFLONE
Keyword(s):  
Cell Surface ◽  
Guinea Pig ◽  
Structure Function ◽  
Receptor Binding ◽  
Core Region ◽  
Terminal Region ◽  
Synthetic Activity ◽  
Triacylglycerol Synthesis ◽  
Acylation Stimulating Protein ◽  
Synthesis Stimulation

Acylation-stimulating protein (ASP or C3adesArg) is a potent lipogenic factor in human and murine adipocytes and fibroblasts. The arginated form of ASP, i.e. complement C3a (C3a), stimulates immunological responses in human granulocytes, mast cells, guinea pig platelets and guinea pig macrophages; however, ASP is inactive in stimulating these responses. Thus both ASP and C3a are bioactive across species but are not functionally interchangeable. Tertiary structure of both proteins by X-ray crystallography and NMR spectroscopy predicts a tightly linked core region consisting of three α-helices linked via three disulphide bonds, with one of the α-helices extending out from the core and terminating in a flexible conformationally irregular carboxy-tail region. The present studies were undertaken in order to define the functionally active domains of ASP, distinctive from those of C3a, using chemical modifications, enzymic cleavage and synthetic peptide fragments. The results indicate that: (i) the N-terminal region (< 10 amino acids) plays little role in ASP receptor binding and triacylglycerol synthesis stimulation; (ii) the native C-terminal region had no activity, but modifications which increased hydrophobicity increased receptor binding, and led to some activation of triacylglycerol synthesis stimulation; (iii) an intact disulphide-linked core region is essential for triacylglycerol synthesis stimulation activity but not for receptor interaction. Finally, basic charges in the carboxy region (His) are essential for ASP triacylglycerol synthesis stimulation but not for receptor binding, whereas both functions are eliminated by the modification of Lys in the disulphide-linked core region. The present results suggest that there are two functional domains in ASP, one that is responsible for the initial binding to the cell surface receptor, and a second domain that activates and increases triacylglycerol synthesis stimulation. This contrasts markedly with the structure-function studies of C3a where both binding competency and function were dependent on the C-terminal Arg. Thus ASP demonstrates distinct bioactivity.

scholarly journals Genetic and Structural Characterization of the Core Region of the Lipopolysaccharide from Serratia marcescens N28b (Serovar O4)

2004 ◽  
Vol 186 (4) ◽  
pp. 978-988 ◽  
Author(s):  
Núria Coderch ◽  
Núria Piqué ◽  
Buko Lindner ◽  
Nihal Abitiu ◽  
Susana Merino ◽  
...  
Keyword(s):  
Serratia Marcescens ◽  
Gene Cluster ◽  
Inner Core ◽  
Core Region ◽  
Complementation Analysis ◽  
Ionization Mass ◽  
Ion Cyclotron Resonance ◽  
The Core ◽  
K 12 ◽  
Ulosonic Acid

ABSTRACT The gene cluster (waa) involved in Serratia marcescens N28b core lipopolysaccharide (LPS) biosynthesis was identified, cloned, and sequenced. Complementation analysis of known waa mutants from Escherichia coli K-12, Salmonella enterica, and Klebsiella pneumoniae led to the identification of five genes coding for products involved in the biosynthesis of a shared inner core structure: [l,d-HeppIIIα(1→7)-l,d-HeppIIα(1→3)-l,d-HeppIα(1→5)-KdopI(4←2)αKdopII] (l,d-Hepp, l-glycero-d-manno-heptopyranose; Kdo, 3-deoxy-d-manno-oct-2-ulosonic acid). Complementation and/or chemical analysis of several nonpolar mutants within the S. marcescens waa gene cluster suggested that in addition, three waa genes were shared by S. marcescens and K. pneumoniae, indicating that the core region of the LPS of S. marcescens and K. pneumoniae possesses additional common features. Chemical and structural analysis of the major oligosaccharide from the core region of LPS of an O-antigen-deficient mutant of S. marcescens N28b as well as complementation analysis led to the following proposed structure: β-Glc-(1→6)-α-Glc-(1→4))-α-d-GlcN-(1→4)-α-d-GalA-[(2←1)-α-d,d-Hep-(2←1)-α-Hep]-(1→3)-α-l,d-Hep[(7←1)-α-l,d-Hep]-(1→3)-α-l,d-Hep-[(4←1)-β-d-Glc]-(1→5)-Kdo. The D configuration of the β-Glc, α-GclN, and α-GalA residues was deduced from genetic data and thus is tentative. Furthermore, other oligosaccharides were identified by ion cyclotron resonance-Fourier-transformed electrospray ionization mass spectrometry, which presumably contained in addition one residue of d-glycero-d-talo-oct-2-ulosonic acid (Ko) or of a hexuronic acid. Several ions were identified that differed from others by a mass of +80 Da, suggesting a nonstoichiometric substitution by a monophosphate residue. However, none of these molecular species could be isolated in substantial amounts and structurally analyzed. On the basis of the structure shown above and the analysis of nonpolar mutants, functions are suggested for the genes involved in core biosynthesis.

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