Jerica Sabotič

• PhD in biotechnology
• Principal Researcher at Jožef Stefan Institute

Listeria monocytogenes is a pathogenic bacterium that can form biofilms in food processing plants, allowing the bacteria to survive despite the control measures applied. As the surface of the bacteria is covered with versatile polysaccharides and proteins, these influence the interactions of the bacterium with any surface. The unique properties and...

The pathogenic bacterium Campylobacter jejuni is a major food safety concern as it can form biofilms that increase its survival and infective potential. Biofilms consist of microbial cells and extracellular matrix (ECM), which is made of water and extracellular polymeric substances (EPS), which are critical for structural integrity and pathogenicit...

Campylobacter jejuni, a Gram-negative bacterium, is one of the most common causes of foodborne illness worldwide. Its adhesion mechanism is mediated by several bacterial factors, including flagellum, protein adhesins, lipooligosaccharides, proteases, and host factors, such as surface glycans on epithelial cells and mucins. Fungal lectins, specializ...

Certain soil insects, such as the root-damaging larvae of the maize pest Diabrotica virgifera virgifera (Coleoptera: Chrysomelidae), are increasingly difficult to control because of recent bans of some insecticides. An alternative and safer approach may be the development of biopesticides based on entomotoxic defense proteins of higher fungi. Many...

Biocatalysis is one of the greatest tools for implementing the 12 principles of Green chemistry. Biocatalysts are bio-based, highly efficient and selective, operate at moderate conditions, and can be reused multiple times. However, the wider application of biocatalysts is plagued by a plethora of drawbacks, such as poor stability at operating condi...

Bioassays are the main tool to decipher bioactivities from natural resources thus their selection and quality are critical for optimal bioprospecting. They are used both in the early stages of compounds isolation/purification/identification, and in later stages to evaluate their safety and efficacy. In this review, we provide a comprehensive overvi...

Cystatin F, a cysteine peptidase inhibitor, is a potent modulator of NK cytotoxicity. By inhibiting granule-mediated cytotoxicity pathway, cystatin F induces formation of non-functional NK cell stage, called split-anergy. We show that N-glycosylation determines the localization and cellular function of cystatin F. Cystatin F mostly exhibited high-m...

Wall teichoic acids (WTA) make up the majority of carbohydrates in the cell wall of Listeria. They play an important physiological role and are involved in host and environmental interactions. Most is known about the WTA of the foodborne pathogen Listeria monocytogenes. Since the non-pathogenic Listeria innocua is its close relative, it has been pr...

The development of antimicrobial resistance and the formation of Salmonella biofilms are serious public health problems. For this reason, new natural compounds with antimicrobial and anti-biofilm activity are being sought, and wild fungi represent an untapped potential. Various extraction agents, including organic solvents and aqueous buffers, can...

Marmorirana smrdljivka (Halyomorpha halys (Stål, 1855); [Hemiptera, Pentatomidae]) je predstavnik družine ščitastih stenic. Gre za invazivno polifagno vrsto, ki izvira iz Vzhodne Azije. Sredi 90-ih je bila vnesena v ZDA ter leta 2004 v Evropo. Danes je prisotna v večini evropskih držav. V zadnjih 15 letih so v gobah odkrili številne entomotoksične...

Background Due to their capacity to specifically recognize subtle alterations in glycoproteins on the cell surface lectins are increasingly being used in diagnostic (for identification of malignant or premalignant cells) and therapeutic purposes (for targeted drug delivery). Materials and Methods We have previously isolated and biochemically chara...

Fungivory of mycorrhizal hyphae has a significant impact on fungal fitness and, by extension, on nutrient transfer between fungi and host plants in natural ecosystems. Mycorrhizal fungi have therefore evolved an arsenal of chemical compounds that are hypothesized to protect the hyphal tissues from being eaten, such as the protease inhibitors mycocy...

We introduce a new family of fungal protease inhibitors with β-trefoil fold from the mushroom Coprinopsis cinerea, named cocaprins, which inhibit both cysteine and aspartic proteases. Two cocaprin-encoding genes are differentially expressed in fungal tissues. One is highly transcribed in vegetative mycelium and the other in the stipes of mature fru...

Clitocybe nebularis lectin (CNL) is a GalNAcβ1‐4GlcNAc‐binding lectin that exhibits an antiproliferative effect exclusively on the Jurkat leukemic T cell line by provoking homotypic aggregation and dose‐dependent cell death. Cell death of Jurkat cells exhibited typical features of early apoptosis, but lacked the activation of initiating and executi...

Biofilms serve as a bacterial survival strategy, allowing bacteria to persist under adverse environmental conditions. The non-pathogenic Listeria innocua is used as a surrogate organism for the foodborne pathogen Listeria monocytogenes, because they share genetic and physiological similarities and can be used in a Biosafety Level 1 laboratory. Seve...

Salmonella spp. are a commonly identified cause of outbreaks of food-borne diseases. Despite much research, there remains the need to find new antimicrobial and anti-biofilm agents against Salmonella. For this, it is necessary to distinguish between these two aspects. Agents that influence biofilm formation should not affect bacterial growth, to th...

Listeria innocua is genetically closely related to the foodborne human pathogen Listeria monocytogenes. However, as most L. innocua strains are non-pathogenic, it has been proposed as a surrogate organism for determining the efficacy of antimicrobial strategies against L. monocytogenes. Teichoic acids are one of the three major cell wall components...

Listeria monocytogenes is a highly pathogenic foodborne bacterium that is ubiquitous in the natural environment and capable of forming persistent biofilms in food processing environments. This species has a rich repertoire of surface structures that enable it to survive, adapt and persist in various environments and promote biofilm formation. We re...

Coastal countries have traditionally relied on the existing marine resources (e.g., fishing, food, transport, recreation, and tourism) as well as tried to support new economic endeavors (ocean energy, desalination for water supply, and seabed mining). Modern societies and lifestyle resulted in an increased demand for dietary diversity, better healt...

Coastal countries have traditionally relied on the existing marine resources (e.g., fishing, food, transport, recreation, and tourism) as well as tried to support new economic endeavors (ocean energy, desalination for water supply, and seabed mining). Modern societies and lifestyle resulted in an increased demand for dietary diversity, better healt...

Studies of biofilm formation by bacteria are crucial for understanding bacterial resistance and for development of novel antibacterial strategies. We have developed a new bioluminescence biofilm assay for Listeria innocua, which is considered a non-pathogenic surrogate for Listeria monocytogenes. L. innocua was transformed with a plasmid for induci...

Lectins have been increasingly utilized as carriers for targeted drug delivery based on their specific binding to glycans located on mammalian cells. This study employed two lectins, B subunit of bacterial Shiga holotoxin (Stx1B) and fungal Clitocybe nebularis lectin (CNL), for surface display on the lactic acid bacterium Lactococcus lactis. The sp...

Cystatin F is a protein inhibitor of cysteine cathepsins, peptidases involved in the activation of the effector molecules of the perforin/granzyme pathway. Cystatin F was previously shown to regulate natural killer cell cytotoxicity. Here, we show that extracellular cystatin F has a role in regulating the killing efficiency of cytotoxic T lymphocyt...

Over the last 2 decades, several coronaviruses (CoVs) have crossed the species barrier into humans, causing highly prevalent and severe respiratory diseases, often with fatal outcomes. CoVs are a large group of enveloped, single-stranded, positive-sense RNA viruses, which encode large replicase polyproteins that are processed by viral peptidases to...

Ralstonia solanaceraum is the quarantine plant pathogenic bacterium that causes bacterial wilt in over 200 host plants, which include economically important crops such as potato, tomato, tobacco, banana, and ginger. Alternative biological methods of disease control that can be used in integrated pest management are extensively studied. In search of...

Marine organisms produce a vast diversity of metabolites with biological activities useful for humans, e.g., cytotoxic, antioxidant, anti-microbial, insecticidal, herbicidal, anticancer, pro-osteogenic and pro-regenerative, analgesic, anti-inflammatory, anti-coagulant, cholesterol-lowering, nutritional, photoprotective, horticultural or other benef...

Clitocybe nebularis lectin (CNL) is present in fruiting bodies of clouded agaric along with several similar isolectins that are all small and stable proteins. It is a beta-trefoil type lectin forming homodimers that are essential for its functionality. It binds specifically N,N′-diacetyllactosediamine (GalNAcβ1-4GlcNAc, LacDiNac) and human blood gr...

Cholinesterases (ChE), the enzymes whose primary function is the hydrolysis of choline esters, are widely expressed throughout the nature. Although they have already been found in plants and microorganisms, including ascomycete fungi, this study is the first report of ChE-like activity in fungi of the phylum Basidiomycota. This activity was detecte...

In-gel cholinesterase-like activity with acetylthiocholine chloride (ACh) as a substrate measured in aqueous extracts from selected species of Basidiomycota. Native polyacrylamide gel run at pH 4 and stained in maleate buffer, pH 6 with 5 mM ACh at room temperature for 2h. Electric eel AChE was used as control (C). Weak activity bands are marked wi...

Predicted putative ChEs from 12 basidiomycetes, 5 ascomycetes and 3 early diverging fungi. (XLSX)

Gene tree of the family of carboxylesterases. The known animal cholinesterase proteins are indicated in red, human non-cholinesterase homologs in blue and fungal homologs in black. The proteins were identified based on the presence of a conserved Pfam domain PF00135. (DOCX)

SDS-PAGE analysis of the proteins exhibiting cholinesterase-like activity with acetylthiocholine chloride (ACh). It was measured in crude extracts from Echinoderma echinaceum (32) and Hygrophoropsis aurantiaca (37). After Native PAGE of the crude extract the ChE activity band was excised, eluted overnight by diffusion from gel pieces and analysed b...

Known animal AChEs. (XLSX)

Background Cystatin F is a protein inhibitor of cysteine peptidases, expressed predominantly in immune cells and localised in endosomal/lysosomal compartments. In cytotoxic immune cells cystatin F inhibits both the major pro-granzyme convertases, cathepsins C and H that activate granzymes, and cathepsin L, that acts as perforin activator. Since per...

The cysteine protease inhibitors, clitocypin and macrocypins, from higher fungi (mycocypins), together with the serine protease inhibitors highly specific for trypsin cospin and cnispin from higher fungi (mycospins), display several characteristics that distinguish them from protease inhibitors from other sources. Their high genetic heterogeneity a...

Intercellular distribution of nutrients and coordination of responses to internal and external cues via endogenous signaling molecules are hallmarks of multicellular organisms. Vegetative mycelia of multicellular fungi are syncytial networks of interconnected hyphae resulting from hyphal tip growth, branching, and fusion. Such mycelia can reach con...

Cysteine cathepsins are lysosomal peptidases involved in the regulation of innate and adaptive immune responses. Among the diverse processes, regulation of granule-dependent cytotoxicity of cytotoxic T-lymphocytes (CTLs) and natural killer (NK) cells during cancer progression has recently gained significant attention. The function of cysteine cathe...

Organophosphate pesticide diazinon is a specific inhibitor of acetylcholinesterase (AChE), which is a common neurotoxicity biomarker in environmental studies. In honeybees, AChE exists in two forms having different physiological roles, one existing as a soluble form and the other as membrane-bound. In most studies AChE activity has been analysed wi...

Cystatin F is a cysteine peptidase inhibitor which, unlike other cystatin family members, is targeted to endosomal/lysosomal compartments. It is synthesized as an inactive disulfide-linked dimer which is then converted to an active monomer by proteolytic cleavage of 15 N-terminal residues. Cystatin F has been suggested to regulate the cytotoxicity...

Mushrooms represent promising sources of novel bioactive compounds and can be applied as innovative strategies to control microbial contamination and infection via the food chain. We characterized aqueous extracts from 21 wild basidiomycete mushrooms and the cultivated oyster mushroom, Pleurotus ostreatus, as putative sources of antimicrobial and a...

l-Amino acid oxidases (LAO) are widely distributed enzymes but those from snake venoms have been studied the most. We describe a method for in-gel detection of LAO activities based on H2O2 detection by a horseradish peroxidase-coupled reaction using o-phenylenediamine. Complex substrates and single l-amino acids were used successfully for screening...

Cystatins comprise a superfamily of evolutionarily related proteins, present in all living organisms, from protozoa to mammals. They act as inhibitors of cysteine peptidases although they can also function independently of their inhibitory function. Cysteine cathepsins are implicated in various physiological and pathological processes. In the immun...

Wild growing mushrooms are a rich source of novel proteins with unique features. We have isolated and characterized trypsin inhibitors from two edible mushrooms, the honey fungus (Armillaria mellea) and the parasol mushroom (Macrolepiota procera), and from the poisonous death cap (Amanita phalloides). The trypsin inhibitors isolated: armespin, macr...

Lectins have been recognized as promising carrier molecules for targeted drug delivery. They specifically bind carbohydrate moieties on cell membranes and trigger cell internalization. Fungal lectin MpL (Macrolepiota procera lectin) does not provoke cancer cell cytotoxicity but is able to bind aminopeptidase N (CD13) and integrin α3β1, two glycopro...

Fungi constitute an enormous unexplored pool of protease inhibitors. Only a handful of fungal protease inhibitors have been exhaustively characterized, but they reveal great versatility and many unique features and novel types of inhibitory mechanisms. Small molecule and protein inhibitors of all catalytic classes of proteases have been identified...

Aminopeptidases, together with other proteases, execute and regulate the total and specifically limited protein breakdown involved in plant physiology, raising the possibility of their involvement in response to drought. We have identified, in leaves of Phaseolus vulgaris L., five aminopeptidases (E.C.3.4.11) whose levels of activity changed when t...

L-amino-acid oxidases (LAO) purified from fungi induce cell death in various mammalian cells including human tumor cell lines. The mechanism, however, remains poorly understood. In this study, we aimed to define a precise mechanism of cell death induced in Jurkat and MCF7 cancer cell lines by ApLAO and CgLAO, LAOs isolated from Amanita phalloides a...

Fruiting bodies or sporocarps of dikaryotic (ascomycetous and basidiomycetous) fungi, commonly referred to as mushrooms, are often rich in entomotoxic and nematotoxic proteins that include lectins and protease inhibitors. These protein toxins are thought to act as effectors of an innate defense system of mushrooms against animal predators including...

Current knowledge of the medicinal properties of Basidiomycetes mushroom species of the genus Clitocybe and of the biological activity of C. nebularis fruiting bodies is reviewed. The main focus is the therapeutic potential of lectins from C. nebularis. Species of the genus Clitocybe, including C. nebularis, have not been traditionally considered a...

In total, 150 protein extracts from 94 different basidiomycete and ascomycete wild mushroom species were tested for antibacterial activity against the quarantine plant-pathogen bacterium Ralstonia solanacearum. In in vitro microtiter plate assays, 15 extracts with moderate to high antibacterial activities were identified: 11 completely inhibited ba...

Fungal ricin B-like lectins and protease inhibitors, mycocypins and mycospins, are important mediators in fungal defence against antagonists and all possess the β-trefoil fold. We demonstrate here that fungal β-trefoil proteins interact with each other, in addition to their apparent targets, and that these interactions modulate their biological act...

Fungi constitute an enormous unexplored pool of protease inhibitors. Only a handful of fungal protease inhibitors have been exhaustively characterized, but they reveal great versatility and many unique features and novel types of inhibitory mechanisms. Small molecule and protein inhibitors of all catalytic classes of proteases have been identified...

Colorado potato beetle (Leptinotarsa decemlineata Say, CPB) is a major potato pest that adapts readily to insecticides. Several types of protease inhibitors have previously been investigated as potential control agents, but with limited success. Recently, cysteine protease inhibitors from parasol mushroom, the macrocypins, were reported to inhibit...

Cysteine cathepsins are lysosomal peptidases involved at different levels in the processes of the innate and adaptive immune responses. Some, such as cathepsins B, L, and H are expressed constitutively in most immune cells. In cells of innate immunity they play a role in cell adhesion and phagocytosis. Other cysteine cathepsins are expressed more s...

The recently identified fungal protease inhibitors cnispin, from Clitocybe nebularis, and cospin, from Coprinopsis cinerea, are both β-trefoil proteins highly specific for trypsin. The reactive site residue of cospin, Arg27, is located on the β2-β3 loop. We show here, that the reactive site residue in cnispin is Lys127, located on the β11-β12 loop....

Proteases and their specific inhibitors are ubiquitously distributed and play a key regulatory role in many biological processes. Gene expression and activity of certain proteases has been shown to increase in Triticum aestivum L. leaves under drought, with a major contribution of cysteine proteases, especially in sensitive wheat varieties. However...

Interactions between fungi and prokaryotes are abundant in many ecological systems. A wide variety of biomolecules regulate such interactions and many of them have found medicinal or biotechnological applications. However, studying a fungal-bacterial system at a cellular level is technically challenging. New microfluidic devices provided a platform...

The unique response of desiccation-tolerant, or resurrection plants, to extreme drought is accompanied by major changes in the protein pool, raising the possibility of the involvement of proteases. We detected and characterized proteases present in their active state in leaf extracts of desiccated Ramonda serbica Panč., a resurrection plant from th...

Unlabelled: Lectins are carbohydrate-binding proteins present in all organisms. Some cytoplasmic lectins from fruiting bodies of dikaryotic fungi are toxic against a variety of parasites and predators. We have isolated, cloned and expressed a novel, single domain lectin from Macrolepiota procera, designated MpL. Determination of the crystal struct...

Controlled protein degradation and activation of proenzymes is required for the growth and development of plants and for them to survive abiotic and biotic stresses. Uncontrolled proteolysis, that is often induced by stress, is however deleterious for plants. Proteases are essential for carrying out and regulating protein breakdown, functions that...

Proteins from higher fungi have attracted interest because of their exceptional characteristics. Macrocypins, cysteine protease inhibitors from the parasol mushroom Macrolepiota procera, were evaluated for their adverse effects and their mode of action on the major potato pest Colorado potato beetle (CPB, Leptinotarsa decemlineata Say). They were s...

An epidemic shift in Hepatitis A virus (HAV) infection has been observed in recent years in rapidly developing countries, with increasing numbers of severe adult cases which has led to renewed interest in vaccination. Our approach in vaccine development uses recombinant expression of the highly immunogenic HAV antigen VP1-P2a in food-grade lactic a...

Protein breakdown by proteases is basic to the plant response to abiotic stresses such as drought. A large number of genes encoding proteases or putative proteases exist in plants. Only a few of those involved in the response to drought have been characterized, and their regulation is poorly understood. We have identified two new subtilases from le...

Abstract Protein protease inhibitors are the tools of nature in controlling proteolytic enzymes. They come in different shapes and sizes. The β-trefoil protease inhibitors that come from plants, first discovered by Kunitz, were later complemented with representatives from higher fungi. They inhibit serine (families S1 and S8) and cysteine proteases...

Proteolytic enzymes (also known as proteases, proteinases or peptidases) offer a wide range of applications. They are routinely used in detergent, leather, food and pharmaceutical industries, as well as in medical and basic research. Therefore, effective isolation procedures are of great importance. The chapter describes the use of recently discove...

Mushrooms are rapidly becoming recognized as a promising source of novel proteins. Several proteins showing unique features have been isolated, including lectins, lignocellulolytic enzymes, protease inhibitors and hydrophobins. They can offer solutions to several medical and biotechnological problems such as microbial drug resistance, low crop yiel...

Lectins are carbohydrate-binding proteins that exert their biological activity by binding to specific cell glycoreceptors. We have expressed CNL, a ricin B-like lectin from the basidiomycete Clitocybe nebularis in Escherichia coli. The recombinant lectin, rCNL, agglutinates human blood group A erythrocytes and is specific for the unique glycan N,N'...

Proteolytic enzymes play essential metabolic and regulatory functions in many biological processes and also offer a wide range of biotechnological applications. Because of their essential roles, their proteolytic activity needs to be tightly regulated. Therefore, small molecules and proteins that inhibit proteases can be versatile tools in the fiel...

Abiotic stresses with a dehydration component (drought, salt, and freezing) involve, as a common feature, increased numbers of inactive proteins – denatured, aggregated or oxidatively damaged. Maintaining proteins in their functional conformation, preventing aggregation of non-native proteins, refolding of denatured proteins to their native conform...

Abiotic stresses with a dehydration component (drought, salt, and freezing) involve, as a common feature, increased numbers of inactive proteins – denatured, aggregated or oxidatively damaged. Maintaining proteins in their functional conformation, preventing aggregation of non-native proteins, refolding of denatured proteins to their native conform...

Cospin (PIC1) from Coprinopsis cinerea is a serine protease inhibitor with biochemical properties similar to those of the previously characterized fungal serine protease inhibitors, cnispin from Clitocybe nebularis and LeSPI from Lentinus edodes, classified in the family I66 of the MEROPS protease inhibitor classification. In particular, it exhibit...

Clitocypin and macrocypin are cysteine protease inhibitors of the mycocypin family which is unique to basidiomycetes. We have established that Clitocybe nebularis and Macrolepiota procera each contain genes for both macrocypin and clitocypin. Both are expressed in M. procera but only clitocypin in C. nebularis. Further analysis of mycocypin express...

Abiotic stresses with a dehydration component (drought, salt, and freezing) involve, as a common feature, increased numbers of inactive proteins – denatured, aggregated or oxidatively damaged. Maintaining proteins in their functional conformation, preventing aggregation of non-native proteins, refolding of denatured proteins to their native conform...

Basidiomycete mushrooms are a rich source of unique substances, including lectins, that could potentially be useful in biotechnology or biomedical applications. Lectins are a group of carbohydrate-binding proteins with diverse biological activities and functions. Here, we demonstrate the presence of a number of lectins in the basidiomycete mushroom...

Fruiting body lectins are ubiquitous in higher fungi and characterized by being synthesized in the cytoplasm and up-regulated during sexual development. The function of these lectins is unclear. A lack of phenotype in sexual development upon inactivation of the respective genes argues against a function in this process. We tested a series of charac...

We have isolated aspartic proteases by affinity chromatography from wild growing basidiomycete Clitocybe nebularis. Pepstatin A sensitive fractions from size exclusion chromatography were subjected to Concanavalin A affinity chromatography. N-terminal sequences of the three bands resolved on SDS-PAGE showed sequence similarity to the A01.018 group...

Mycocypins, clitocypins and macrocypins, are cysteine protease inhibitors isolated from the mushrooms Clitocybe nebularis and Macrolepiota procera. Lack of sequence homology to other families of protease inhibitors suggested that mycocypins inhibit their target cysteine protease by a unique mechanism and that a novel fold may be found. The crystal...

A new family of cysteine protease inhibitors from the basidiomycete Macrolepiota procera has been identified and the family members have been termed macrocypins. These macrocypins are encoded by a family of genes that is divided into five groups with more than 90% within-group sequence identity and 75-86% between-group sequence identity. Several di...

We have isolated serine protease inhibitors from the basidiomycete Clitocybe nebularis, CnSPIs, using trypsin affinity chromatography. Full-length gene and cDNA sequences were determined for one of them, named cnispin, and the recombinant protein was expressed in Escherichia coli at high yield. The primary structure and biochemical properties of cn...