Hydrophobins: Exceptional proteins for many applications in brewery environment and other bio-industries
Introduction
Hydrophobins are a large family of small cysteine rich proteins (about 100 amino acids) (Chaplin and Kennedy, 1994) with a molecular mass of around 10 kDa (Cooper and Kennedy, 2010) (Fig. 1). The name hydrophobin was originally used due to their high content of hydrophobic amino acids (Armenante, 2008). The proteins are secreted by fungi into liquid media or remain at the surface of aerial mycelia (Stubner et al., 2010). Recent evidence demonstrates that a broad spectrum of functions in fungal growth and development is mediated by hydrophobins (Armenante, 2008). They have been detected in fungal plant pathogens (Talbot et al., 1996, Bidochka et al., 2001), insect pathogens (St Leger et al., 1992), and human pathogens (Parta et al., 1994) and saprophytes (Bell-Pedersen et al., 1996). More than one type of hydrophobin has been found in most moulds. For example, in Schizophyllum commune, at least four hydrophobin-encoding genes (Sc1–Sc4) with dissimilar functions have been isolated (Wessels, 1993). Due to the distribution of the cysteines and the clustering of hydrophobic and hydrophilic residues, hydrophobins are divided into two classes: class I such as SC3 and EAS, and class II like HFBI and HFBII. The aggregates formed by both classes are distinguished on the basis of their solubility and morphology (Linder, 2009a, Linder, 2009b).
Hydrophobins can assemble at a hydrophilic–hydrophobic interface forming an amphipathic film (Wessels, 2000) (Fig. 2). This film makes hydrophobic surfaces of liquid or solid materials wettable, while hydrophilic surfaces are becoming hydrophobic. Thanks to their extraordinary properties, hydrophobins were consequently first suggested for a number of applications involving the modification of surfaces properties (Janssen et al., 2002a, Janssen et al., 2002b) leading to improving biocompatibility, reducing friction, or providing specific sites for protein immobilization (Kisko, 2008) (Table 1). The self-assembly of hydrophobins (Wösten and Wessels, 1997) is interesting in the field of surfactants, emulsifiers, and surface coating. Using the hydrophobin in future nanotechnology applications (e.g. patterning molecules at a surface with nanometer accuracy) (Scholtmeijer et al., 2001, Wang et al., 2004, Thomas, 1995), may lead to improving biosensors (Bilewicz et al., 2001), and applications in tissue engineering (Lumsdon et al., 2005). Remarkable is the possibility to separate and isolate nutritional proteins of interest by fusing them to the hydrophobins. This could result in a generic, rapid and efficient protein purification system (Linder et al., 2004). As hydrophobins are adsorbed easily to both hydrophilic and hydrophobic surfaces, such as glass, mica, parafilm, and Teflon, resulting in modification of their properties (Wösten et al., 1994) and the ability of hydrophobins to assemble into strong monolayers and to modify the hydrophobicity of a surface these proteins may have a wide range of potential usages as biotechnological tools (Mackay et al., 2001). Increasing demands for hydrophobins leads to a challenge in terms of the production and purification of this product (Subkowski et al., 2007, Winterburn et al., 2011a, Winterburn et al., 2011b).
For the stabilization of oil vesicles, or oil emulsions, in pharmaceutical applications hydrophobins might be used instead of traditional surfactants (Scholtmeijer et al., 2001). Hydrophobins with suitable properties are potential candidates for native biosurfactants to treat repellency. On the other hand, understanding their involvement in modification of hydrophobicity could open up new applications, for instance in using fungal field inoculants (Rillig, 2005). The amphiphilic nature of hydrophobins can also be exploited using its surfactant-like behavior in solutions, since they are quite soluble in water (Lahtinen et al., 2008). Recently, the ability of hydrophobins to increase the dispersion of hydrophobic particles in water has been described (Wang et al., 2010a, Wang et al., 2010b, Wang et al., 2010c).
Although various applications are expected for native hydrophobins, the modification of hydrophobin structures might offer more possibilities as it may lead to changing the biophysical properties of the hydrophobin surface. By changing the surface hydrophobicity, the binding of various molecules and cells can be manipulated (Hektor and Scholtmeijer, 2005). The self-assembly at hydrophobic/hydrophilic interfaces as well as the nontoxic nature of hydrophobins allow them to be introduced into a number of products (Kershaw and Talbot, 1998). A negative aspect of hydrophobins is that they are produced by fungi when growing on grains (e.g. barley) and during grain treatment. Barley is a very important vegetal food with high economic value as it is used in the production of beer and other drinks (Ehrenbergerova et al., 2006, Jood and Singh, 2001, Sehgal and Kawatra, 1998). The presence of hydrophobins leads to gushing, which has many economic drawbacks for brewing industries. Consequently, improvement in rapid and early detection methods for hydrophobins is required (Shokribousjein et al., 2011).
The aim of this report is to emphasize the different applications of hydrophobins and to stimulate more studies for finding methods to detect, isolate and produce these valuable proteins.
Section snippets
Hydrophobins: novel biosurfactants
Finding harmless surface active components is a challenging topic in food industries and hydrophobins have become outstanding candidates in that context (Murray et al., 2009). These unusual proteins seem to adsorb and act as rigid hydrophobic nanoparticles.
Hydrophobins are surfactive molecules that can polymerize at an air/water or water/oil interface and therefore, form a strong, polymeric monolayer which can reverse the hydropathicity of surfaces (Mackay et al., 2001). Their surface activity
Hydrophobins in foam stabilization and gushing
The presence of hydrophobins causes foaming in shaken microbial cultures due to their surface activity. In particular, aerated bioreactor cultures of filamentous fungi often suffer from foaming, and antifoam agents are needed (Linder et al., 2005). The mechanism of foam stabilization by hydrophobins may result in high surface elasticity. Bailey et al. (2002) have studied the problem using T. reesei strains in which the hfb1 or hfb2 genes were either inactivated or overexpressed. Overexpression
Hydrophobins in protein immobilization and separation
Based on the efficient assembly, hydrophobins are used as a tag to purify proteins from complex mixtures. Protein immobilization is an interesting topic in food and nutrition science (Wösten, 2001). The captured proteins might be enzymes or nutrient ingredients. In principle, both class I and class II hydrophobins can be used to immobilize proteins, however, class I hydrophobins are preferred when detergents or pressure are involved.
Class I hydrophobin from Pleurotus ostreatus was used to coat
Hydrophobins for biosensors improvement
Although biosensors are used to detect and/or to determine many compounds, such as food ingredients, or several other phenomena, it appears that biosensors with more sensitivity and more accuracy are needed.
A general problem of biosensors using immobilized enzymes is the denaturation of the proteins bound on surfaces (Chakarova and Carlson, 2004). It has been confirmed that a hydrophobin coating can inhibit denaturation of immobilized enzymes and preserve long time activity. Enzymes can be
Hydrophobins: medical and pharmaceutical applications
Hydrophobins are interesting for enhancing the biocompatibility of medical implants in order to avoid the rejection of the implants (Hektor and Scholtmeijer, 2005). Low-friction surfaces are required in various biomedical applications (Misra et al., 2006). To reduce the friction of materials, hydrophobins can be used safely. It has been suggested (Wösten, 2001) and later confirmed (Aimanianda et al., 2009) that by covering fungal aerial structures, hydrophobins shield antigens in the cell wall,
Hydrophobins: multiple promising applications
There are several possible applications of hydrophobins that were not extensively considered so far. Hydrophobins were found to extend the residence time of hair care products and shampoos (Vic, 2003). They can be used to stabilize emulsions in creams and ointments (Wösten et al., 1994). To develop the stability and particle size of suspensions and emulsions, and to conserve the activity of proteins at surface of a liquid or a solid material, the amphipathic hydrophobins can be exploited (
Conclusion
From what we reported above it can be concluded that the extraordinary properties of hydrophobins offer numerous possibilities for applications in science and technology. Excellent applications are possible in domains necessitating a better stabilization of foams. Related to excessive foaming of carbonated solutions, hydrophobins are excellent indicators for the gushing phenomenon, which is really a serious problem for carbonated beverage industries. For the purification of proteins, fusion
Acknowledgment
This work is supported by the Hydrophobin Chair granted to the KU-Leuven by the Duvel-Moortgat Brewery, the trappist beer breweries Orval and Chimay and Cargill Malting.
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Extraction and spray drying of Class II hydrophobin HFBI produced by Trichoderma reesei
2019, Process BiochemistryCitation Excerpt :Hydrophobins, produced by filamentous fungi, are relatively small (7–10 kDa) surface-active proteins which self-assemble at hydrophobic-hydrophilic interfaces [1].
HFB7 – A novel orphan hydrophobin of the Harzianum and Virens clades of Trichoderma, is involved in response to biotic and abiotic stresses
2017, Fungal Genetics and BiologyCitation Excerpt :Moreover, they are required for the efficient spore dispersion through air, for the attachment to hydrophobic and hydrophilic surfaces and for the stabilization of air channels in fruiting bodies (Wösten et al., 1994; Lugones et al., 1999; Wösten, 2001). Because of their intrinsic property of self-assembly, hydrophobins are potential candidates for a diversity of applications ranging from bulk (foam stabilizers, emulsifiers) to high-value products (biosensors, fusion tags for efficient purification and immobilization, biomedical devices) (Corvis et al., 2005; Janssen et al., 2002; Khalesi et al., 2012; von Vacano et al., 2011). Several studies have also demonstrated hydrophobins as powerful implements in nanoparticle based drug delivery (Bimbo et al., 2011; Fang et al., 2014; Valo et al., 2010; Zhao et al., 2016).
Antioxidant activity and ACE-inhibitory of Class II hydrophobin from wild strain Trichoderma reesei
2016, International Journal of Biological MacromoleculesTechnological possibilities to prevent and suppress primary gushing of beer
2016, Trends in Food Science and TechnologyCitation Excerpt :The authors suggested that ocimene blocked the interaction between the CO2 and hydrophobins. Hydrophobins dispose with one or more hydrophobic patches, which are made of amino acids residues (Khalesi et al., 2012). This hydrophobic patch is responsible for the strong attraction between hydrophobins and CO2.
Fungal biofilm reactor improves the productivity of hydrophobin HFBII
2014, Biochemical Engineering JournalCitation Excerpt :Hydrophobins exhibit a negative effect when present in carbonated beverages contaminated by molds, leading to gushing [5,6]. Nevertheless, due to its outstanding surface-active properties, several positive potential applications such as antifouling agent, drugs formulation, and stabilization of emulsions have been proposed for this family [7]. Class II hydrophobins HFBI and HFBII produced by T. reesei have been structurally studied by many authors [8,9].