Hyperpolarisation of Mitochondrial Membranes Is a Critical Component of the Antifungal Mechanism of the Plant Defensin, Ppdef1.

Plant defensins are a large family of small cationic proteins with diverse functions and mechanisms of action, most of which assert antifungal activity against a broad spectrum of fungi. The partial mechanism of action has been resolved for a small number of members of plant defensins, and studies have revealed that many act by more than one mechanism. The plant defensin Ppdef1 has a unique sequence and long loop 5 with fungicidal activity against a range of human fungal pathogens, but little is known about its mechanism of action.

The Plant Defensin Ppdef1 Is a Novel Topical Treatment for Onychomycosis.

Onychomycosis, or fungal nail infection, causes not only pain and discomfort but can also have psychological and social consequences for the patient. Treatment of onychomycosis is complicated by the location of the infection under the nail plate, meaning that antifungal molecules must either penetrate the nail or be applied systemically. Currently, available treatments are limited by their poor nail penetration for topical products or their potential toxicity for systemic products.

Membrane binding properties of plant defensins.

The membrane interaction characteristics of five antifungal plant defensin peptides: NaD1, and the related HXP4 and L5, as well as NaD2 and the related ZmD32 were studied. These peptides were chosen to cover a broad range of cationic charges with little structural variations, allowing for assessment of the role of charge in their membrane interactions. Membrane permeabilizing activity against C.

Improving the Digestibility of Plant Defensins to Meet Regulatory Requirements for Transgene Products in Crop Protection.

Despite the use of chemical fungicides, fungal diseases have a major impact on the yield and quality of plant produce globally and hence there is a need for new approaches for disease control. Several groups have examined the potential use of antifungal plant defensins for plant protection and have produced transgenic plants expressing plant defensins with enhanced resistance to fungal disease. However, before they can be developed commercially, transgenic plants must pass a series of strict regulations to ensure that they are safe for human and animal consumption as well as the environment.

Screening the Nonessential Gene Deletion Library Reveals Diverse Mechanisms of Action for Antifungal Plant Defensins.

Plant defensins are a large family of proteins, most of which have antifungal activity against a broad spectrum of fungi. However, little is known about how they exert their activity. The mechanisms of action of only a few members of the family have been investigated and, in most cases, there are still a number of unknowns.

A suite of kinetically superior AEP ligases can cyclise an intrinsically disordered protein.

Asparaginyl endopeptidases (AEPs) are a class of enzymes commonly associated with proteolysis in the maturation of seed storage proteins. However, a subset of AEPs work preferentially as peptide ligases, coupling release of a leaving group to formation of a new peptide bond. These "ligase-type" AEPs require only short recognition motifs to ligate a range of targets, making them useful tools in peptide and protein engineering for cyclisation of peptides or ligation of separate peptides into larger products.

The interaction with fungal cell wall polysaccharides determines the salt tolerance of antifungal plant defensins.

The fungal cell wall is the first point of contact between fungal pathogens and host organisms. It serves as a protective barrier against biotic and abiotic stresses and as a signal to the host that a fungal pathogen is present. The fungal cell wall is made predominantly of carbohydrates and glycoproteins, many of which serve as binding receptors for host defence molecules or activate host immune responses through interactions with membrane-bound receptors.

Salt-Tolerant Antifungal and Antibacterial Activities of the Corn Defensin ZmD32.

Pathogenic microbes are developing resistance to established antibiotics, making the development of novel antimicrobial molecules paramount. One major resource for discovery of antimicrobials is the arsenal of innate immunity molecules that are part of the first line of pathogen defense in many organisms. Gene encoded cationic antimicrobial peptides are a major constituent of innate immune arsenals.

The evolution, function and mechanisms of action for plant defensins.

Plant defensins are an extensive family of small cysteine rich proteins characterised by a conserved cysteine stabilised alpha beta protein fold which resembles the structure of insect and vertebrate defensins. However, secondary structure and disulphide topology indicates two independent superfamilies of defensins with similar structures that have arisen via an extreme case of convergent evolution. Defensins from plants and insects belong to the cis-defensin superfamily whereas mammalian defensins belong to the trans-defensin superfamily.

The Plant Defensin NaD1 Enters the Cytoplasm of Candida Albicans via Endocytosis.

Antimicrobial peptides are widespread in nature and are produced by many organisms as a first line of defence against pathogens. These peptides have a broad range of biological activities, such as antibacterial or antifungal activities and act with varied mechanisms of action. A large number of the peptides are amphipathic α-helices which act by disrupting plasma membranes and allowing leakage of intracellular contents.

Co-expression of a cyclizing asparaginyl endopeptidase enables efficient production of cyclic peptides in planta.

Cyclotides are ultra-stable, backbone-cyclized plant defence peptides that have attracted considerable interest in the pharmaceutical industry. This is due to their range of native bioactivities as well as their ability to stabilize other bioactive peptides within their framework. However, a hindrance to their widespread application is the lack of scalable, cost-effective production strategies.

Synergistic Activity between Two Antifungal Proteins, the Plant Defensin NaD1 and the Bovine Pancreatic Trypsin Inhibitor.

Defensins are a large family of small, cationic, cysteine-rich proteins that are part of the defense arsenal that plants use for protection against potentially damaging fungal infections. The plant defensin NaD1 from is a potent antifungal protein that inhibits growth and kills a variety of fungal pathogens that affect both plant and animal (human) hosts. Some serine protease inhibitors have also been reported to be antifungal molecules, while others have no inhibitory activity against fungi.

Nicotiana alata Defensin Chimeras Reveal Differences in the Mechanism of Fungal and Tumor Cell Killing and an Enhanced Antifungal Variant.

The plant defensin NaD1 is a potent antifungal molecule that also targets tumor cells with a high efficiency. We examined the features of NaD1 that contribute to these two activities by producing a series of chimeras with NaD2, a defensin that has relatively poor activity against fungi and no activity against tumor cells. All plant defensins have a common tertiary structure known as a cysteine-stabilized α-β motif which consists of an α helix and a triple-stranded β-sheet stabilized by four disulfide bonds.

The plant defensin NaD1 introduces membrane disorder through a specific interaction with the lipid, phosphatidylinositol 4,5 bisphosphate.

Plant defensins interact with phospholipids in bilayers as part of their cytotoxic activity. Solanaceous class II defensins with the loop 5 sequence pattern "S-[KR]-[ILVQ]-[ILVQ]-[KR]-[KR]" interact with PI(4,5)P2. Here, the prototypical defensin of this class, NaD1, is used to characterise the biophysical interactions between these defensins and phospholipid bilayers.

Structural homology guided alignment of cysteine rich proteins.

Background: Cysteine rich protein families are notoriously difficult to align due to low sequence identity and frequent insertions and deletions.

Results: Here we present an alignment method that ensures homologous cysteines align by assigning a unique 10 amino acid barcode to those identified as structurally homologous by the DALI webserver. The free inter-cysteine regions of the barcoded sequences can then be aligned using any standard algorithm.

Efficient backbone cyclization of linear peptides by a recombinant asparaginyl endopeptidase.

Cyclotides are diverse plant backbone cyclized peptides that have attracted interest as pharmaceutical scaffolds, but fundamentals of their biosynthetic origin remain elusive. Backbone cyclization is a key enzyme-mediated step of cyclotide biosynthesis and confers a measure of stability on the resultant cyclotide. Furthermore, cyclization would be desirable for engineered peptides.

Bovine pancreatic trypsin inhibitor is a new antifungal peptide that inhibits cellular magnesium uptake.

Antimicrobial peptides (AMPs) are promising agents for control of bacterial and fungal infections. Traditionally, AMPs were thought to act through membrane disruption but recent experiments have revealed a diversity of mechanisms. Here we describe a novel antifungal activity for bovine pancreatic trypsin inhibitor (BPTI).

Phosphoinositide-mediated oligomerization of a defensin induces cell lysis.

Cationic antimicrobial peptides (CAPs) such as defensins are ubiquitously found innate immune molecules that often exhibit broad activity against microbial pathogens and mammalian tumor cells. Many CAPs act at the plasma membrane of cells leading to membrane destabilization and permeabilization. In this study, we describe a novel cell lysis mechanism for fungal and tumor cells by the plant defensin NaD1 that acts via direct binding to the plasma membrane phospholipid phosphatidylinositol 4,5-bisphosphate (PIP2).

Agp2p, the plasma membrane transregulator of polyamine uptake, regulates the antifungal activities of the plant defensin NaD1 and other cationic peptides.

Cationic antifungal peptides (AFPs) act through a variety of mechanisms but share the common feature of interacting with the fungal cell surface. NaD1, a defensin from Nicotiana alata, has potent antifungal activity against a variety of fungi of both hyphal and yeast morphologies. The mechanism of action of NaD1 occurs via three steps: binding to the fungal cell surface, permeabilization of the plasma membrane, and internalization and interaction with intracellular targets to induce fungal cell death.

Activation of stress signalling pathways enhances tolerance of fungi to chemical fungicides and antifungal proteins.

Fungal disease is an increasing problem in both agriculture and human health. Treatment of human fungal disease involves the use of chemical fungicides, which generally target the integrity of the fungal plasma membrane or cell wall. Chemical fungicides used for the treatment of plant disease, have more diverse mechanisms of action including inhibition of sterol biosynthesis, microtubule assembly and the mitochondrial respiratory chain.

Identification and mechanism of action of the plant defensin NaD1 as a new member of the antifungal drug arsenal against Candida albicans.

In recent decades, pathogenic fungi have become a serious threat to human health, leading to major efforts aimed at characterizing new agents for improved treatments. Promising in this context are antimicrobial peptides produced by animals and plants as part of innate immune systems. Here, we describe an antifungal defensin, NaD1, with activity against the major human pathogen Candida albicans, characterize the mechanism of killing, and identify protection strategies used by the fungus to survive defensin treatment.

A radish seed antifungal peptide with a high amyloid fibril-forming propensity.

The amyloid fibril-forming ability of two closely related antifungal and antimicrobial peptides derived from plant defensin proteins has been investigated. As assessed by sequence analysis, thioflavin T binding, transmission electron microscopy, atomic force microscopy and X-ray fiber diffraction, a 19 amino acid fragment from the C-terminal region of Raphanus sativus antifungal protein, known as RsAFP-19, is highly amyloidogenic. Further, its fibrillar morphology can be altered by externally controlled conditions.