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Margaret Sunde - One of the best experts on this subject based on the ideXlab platform.
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Cell-free expression of natively folded Hydrophobins.
Protein expression and purification, 2020Co-Authors: Rezwan Siddiquee, Margaret Sunde, Samuel Sung-chan Choi, Shirley Siuley Lam, Patrick Wang, Gottfried Otting, Ann Hau-yu KwanAbstract:Abstract Hydrophobins are a family of cysteine-rich proteins unique to filamentous fungi. The proteins are produced in a soluble form but self-assemble into organised amphipathic layers at hydrophilic:hydrophobic interfaces. These layers contribute to transitions between wet and dry environments, spore dispersal and attachment to surfaces for growth and infection. Hydrophobins are characterised by four disulphide bonds that are critical to their structure and function. Thus, obtaining correctly folded, soluble and functional Hydrophobins directly from bacterial recombinant expression is challenging and in most cases, initial denaturation from inclusion bodies followed by oxidative refolding are required to obtain folded proteins. Here, we report the use of cell-free expression with E. coli cell lysate to directly obtain natively folded Hydrophobins. All six of the Hydrophobins tested could be expressed after optimisation of redox conditions. For some Hydrophobins, the inclusion of the disulfide isomerase DsbC further enhanced expression levels. We are able to achieve a yield of up to 1 mg of natively folded hydrophobin per mL of reaction. This has allowed the confirmation of the correct folding of Hydrophobins with the use of 15N-cysteine and 15N–1H nuclear magnetic resonance experiments within 24 h of starting from plasmid stocks.
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Fungal Hydrophobins and Their Self-Assembly into Functional Nanomaterials.
Advances in experimental medicine and biology, 2019Co-Authors: Jennifer I-chun Lai, Margaret SundeAbstract:In recent years, much attention has focused on incorporating biological and bio-inspired nanomaterials into various applications that range from functionalising surfaces and enhancing biomolecule binding properties, to coating drugs for improved bioavailability and delivery. Hydrophobin proteins, which can spontaneously assemble into amphipathic layers at hydrophobic:hydrophilic interfaces, are exciting candidates for use as nanomaterials. These unique proteins, which are only expressed by filamentous fungi, have been the focus of increasing interest from the biotechnology industry, as evidenced by the sharply growing number of hydrophobin-associated publications and patents. Here, we explore the contribution of different Hydrophobins to supporting fungal growth and development. We describe the key structural elements of Hydrophobins and the molecular characteristics that underlie self-assembly of these proteins at interfaces. We outline the multiple roles that Hydrophobins can play in supporting aerial growth of filamentous structures, facilitating spore dispersal and preventing an immune response in the infected host. The growing understanding of the hydrophobin protein structure and self-assembly process highlights the potential for hydrophobin proteins to be engineered for use in a variety of novel applications that require biocompatible coatings.
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Fungal Hydrophobin Proteins Produce Self-Assembling Protein Films with Diverse Structure and Chemical Stability.
Nanomaterials (Basel Switzerland), 2014Co-Authors: Qin Ren, Ann H. Kwan, Chi L.l. Pham, Vanessa K. Morris, Margaret SundeAbstract:Hydrophobins are small proteins secreted by fungi and which spontaneously assemble into amphipathic layers at hydrophilic-hydrophobic interfaces. We have examined the self-assembly of the Class I Hydrophobins EAS∆15 and DewA, the Class II hydrophobin NC2 and an engineered chimeric hydrophobin. These Class I Hydrophobins form layers composed of laterally associated fibrils with an underlying amyloid structure. These two Class I Hydrophobins, despite showing significant conformational differences in solution, self-assemble to form fibrillar layers with very similar structures and require a hydrophilic-hydrophobic interface to trigger self-assembly. Addition of additives that influence surface tension can be used to manipulate the fine structure of the protein films. The Class II hydrophobin NC2 forms a mesh-like protein network and the engineered chimeric hydrophobin displays two multimeric forms, depending on assembly conditions. When formed on a graphite surface, the fibrillar EAS∆15 layers are resistant to alcohol, acid and basic washes. In contrast, the NC2 Class II monolayers are dissociated by alcohol treatment but are relatively stable towards acid and base washes. The engineered chimeric Class I/II hydrophobin shows increased stability towards alcohol and acid and base washes. Self-assembled hydrophobin films may have extensive applications in biotechnology where biocompatible; amphipathic coatings facilitate the functionalization of nanomaterials.
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Backbone and sidechain ^1H, ^13C and ^15N chemical shift assignments of the hydrophobin MPG1 from the rice blast fungus Magnaporthe oryzae
Biomolecular NMR Assignments, 2013Co-Authors: Anthony A. Rey, Antoine Hocher, Ann H. Kwan, Margaret SundeAbstract:Fungal Hydrophobins are secreted proteins that self-assemble at hydrophobic:hydrophilic interfaces. They are essential for a variety of processes in the fungal life cycle, including mediating interactions with surfaces and infection of hosts. The fungus Magnaporthe oryzae , the causative agent of rice blast, relies on the unique properties of Hydrophobins to infect cultivated rice as well as over 50 different grass species. The hydrophobin MPG1 is highly expressed during rice blast pathogenesis and has been implicated during host infection. Here we report the backbone and sidechain assignments for the class I hydrophobin MPG1 from the rice blast fungus Magnaporthe oryzae .
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analysis of the structure and conformational states of dewa gives insight into the assembly of the fungal Hydrophobins
Journal of Molecular Biology, 2013Co-Authors: Vanessa K. Morris, Ann H. Kwan, Margaret SundeAbstract:The hydrophobin DewA from the fungus Aspergillus nidulans is a highly surface-active protein that spontaneously self-assembles into amphipathic monolayers at hydrophobic:hydrophilic interfaces. These monolayers are composed of fibrils that are a form of functional amyloid. While there has been significant interest in the use of DewA for a variety of surface coatings and as an emulsifier in biotechnological applications, little is understood about the structure of the protein or the mechanism of self-assembly. We have solved the solution NMR structure of DewA. While the pattern of four disulfide bonds that is a defining feature of Hydrophobins is conserved, the arrangement and composition of secondary-structure elements in DewA are quite different to what has been observed in other hydrophobin structures. In addition, we demonstrate that DewA populates two conformations in solution, both of which are assembly competent. One conformer forms a dimer at high concentrations, but this dimer is off-pathway to fibril formation and may represent an assembly control mechanism. These data highlight the structural differences between fibril-forming Hydrophobins and those that form amorphous monolayers. This work will open up new opportunities for the engineering of Hydrophobins with novel biotechnological applications.
Markus Linder - One of the best experts on this subject based on the ideXlab platform.
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The dynamics of multimer formation of the amphiphilic hydrophobin protein HFBII.
Colloids and surfaces. B Biointerfaces, 2017Co-Authors: Mathias S. Grunér, Arja Paananen, Géza R. Szilvay, Markus LinderAbstract:Hydrophobins are surface-active proteins produced by filamentous fungi. They have amphiphilic structures and form multimers in aqueous solution to shield their hydrophobic regions. The proteins rearrange at interfaces and self-assemble into films that can show a very high degree of structural order. Little is known on dynamics of multimer interactions in solution and how this is affected by other components. In this work we examine the multimer dynamics by stopped-flow fluorescence measurements and Forster Resonance Energy Transfer (FRET) using the class II hydrophobin HFBII. The half-life of exchange in the multimer state was 0.9s at 22°C with an activation energy of 92kJ/mol. The multimer exchange process of HFBII was shown to be significantly affected by the closely related HFBI hydrophobin, lowering both activation energy and half-life for exchange. Lower molecular weight surfactants interacted in very selective ways, but other surface active proteins did not influence the rates of exchange. The results indicate that the multimer formation is driven by specific molecular interactions that distinguish different Hydrophobins from each other.
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Novel Hydrophobin Fusion Tags for Plant-Produced Fusion Proteins
PloS one, 2016Co-Authors: Lauri Reuter, Markus Linder, Anneli Ritala, Jussi JoensuuAbstract:Hydrophobin fusion technology has been applied in the expression of several recombinant proteins in plants. Until now, the technology has relied exclusively on the Trichoderma reesei hydrophobin HFBI. We screened eight novel hydrophobin tags, T. reesei HFBII, HFBIII, HFBIV, HFBV, HFBVI and Fusarium verticillioides derived HYD3, HYD4 and HYD5, for production of fusion proteins in plants and purification by two-phase separation. To study the properties of the Hydrophobins, we used N-terminal and C-terminal GFP as a fusion partner. Transient expression of the hydrophobin fusions in Nicotiana benthamiana revealed large variability in accumulation levels, which was also reflected in formation of protein bodies. In two-phase separations, only HFBII and HFBIV were able to concentrate GFP into the surfactant phase from a plant extract. The separation efficiency of both tags was comparable to HFBI. When the accumulation was tested side by side, HFBII-GFP gave a better yield than HFBI-GFP, while the yield of HFBIV-GFP remained lower. Thus we present here two alternatives for HFBI as functional fusion tags for plant-based protein production and first step purification.
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Charge-based engineering of hydrophobin HFBI: effect on interfacial assembly and interactions.
Biomacromolecules, 2015Co-Authors: Michael Lienemann, Arja Paananen, Mathias S. Grunér, Matti Siika-aho, Markus LinderAbstract:Hydrophobins are extracellular proteins produced by filamentous fungi. They show a variety of functions at interfaces that help fungi to adapt to their environment by, for example, adhesion, formation of coatings, and lowering the surface tension of water. Hydrophobins fold into a globular structure and have a distinct hydrophobic patch on their surface that makes these proteins amphiphilic. Their amphiphilicity implies interfacial assembly, but observations indicate that intermolecular interactions also contribute to their functional properties. Here, we used the class II hydrophobin HFBI from Trichoderma reesei as a model to understand the structural basis for the function of Hydrophobins. Four different variants were made in which charged residues were mutated. The residues were chosen to probe the role of different regions of the hydrophilic part of the proteins. Effects of the mutations were studied by analyzing the formation and structure of self-assembled layers, multimerization in solution, surfac...
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Hydrophobins as aqueous lubricant additive for a soft sliding contact
Colloids and Surfaces B: Biointerfaces, 2014Co-Authors: Kirsi I. Pakkanen, Markus LinderAbstract:Abstract Two type II fungal Hydrophobins, HFBI and FpHYD5, have been studied as aqueous lubricant additive at a nonpolar, compliant sliding contact (self-mated poly(dimethylsiloxane) (PDMS) contact) at two different concentrations, 0.1 mg/mL and 1.0 mg/mL. The two Hydrophobins are featured as non-glycosylated (HFBI, m.w. ca. 7 kDa) vs glycosylated (FpHYD5, m.w. ca. 10 kDa) proteins. Far UV CD spectra of the two Hydrophobins were very similar, suggesting overall structural similarity, but showed a noticeable difference according to the concentration. This is proposed to be related to the formation of multimers at 1.0 mg/mL. Despite 10-fold difference in the bulk concentration, the adsorbed masses of the Hydrophobins onto PDMS surface obtained from the two solutions (0.1 and 1.0 mg/mL) were nearly identical, suggesting that a monolayer of the Hydrophobins are formed from 0.1 mg/mL solution. PDMS–PDMS sliding interface was effectively lubricated by the hydrophobin solutions, and showed a reduction in the coefficient of friction by as much as ca. two orders of magnitude. Higher concentration solution (1.0 mg/mL) provided a superior lubrication, particularly in low-speed regime, where boundary lubrication characteristic is dominant via ‘self-healing’ mechanism. FpHYD5 revealed a better lubrication than HFBI presumably due to the presence of glycans and improved hydration of the sliding interface. Two type II Hydrophobins function more favorably compared to a synthetic amphiphilic copolymer, PEO–PPO–PEO, with a similar molecular weight. This is ascribed to higher amount of adsorption of the Hydrophobins to hydrophobic surfaces from aqueous solution.
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Structure-function relationships in Hydrophobins: Probing the role of charged side chains
Applied and environmental microbiology, 2013Co-Authors: Michael Lienemann, Julieanne Gandier, Emma R Master, Maija Tenkanen, Jussi Joensuu, Atsushi Iwanaga, Yoshiyuki Takatsuji, Tetsuya Haruyama, Markus LinderAbstract:Hydrophobins are small fungal proteins that are amphiphilic and have a strong tendency to assemble at interfaces. By taking advantage of this property, Hydrophobins have been used for a number of applications: as affinity tags in protein purification, for protein immobilization, such as in foam stabilizers, and as dispersion agents for insoluble drug molecules. Here, we used site-directed mutagenesis to gain an understanding of the molecular basis of their properties. We especially focused on the role of charged amino acids in the structure of Hydrophobins. For this purpose, fusion proteins consisting of Trichoderma reesei hydrophobin I (HFBI) and the green fluorescent protein (GFP) that contained various combinations of substitutions of charged amino acids (D30, K32, D40, D43, R45, K50) in the HFBI structure were produced. The effects of the introduced mutations on binding, oligomerization, and partitioning were characterized in an aqueous two-phase system. It was found that some substitutions caused better surface binding and reduced oligomerization, while some showed the opposite effects. However, all mutations decreased partitioning in surfactant systems, indicating that the different functions are not directly correlated and that partitioning is dependent on finely tuned properties of Hydrophobins. This work shows that not all functions in self-assembly are connected in a predictable way and that a simple surfactant model for hydrophobin function is insufficient.
Francis Martin - One of the best experts on this subject based on the ideXlab platform.
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The hydrophobin-like OmSSP1 may be an effector in the ericoid mycorrhizal symbiosis
Frontiers in plant science, 2018Co-Authors: Salvatore Casarrubia, Francis Martin, Annegret Kohler, Stefania Daghino, Emmanuelle Morin, Hassine Radhouane Khouja, Yohann Daguerre, Claire Veneault-fourrey, Silvia Perotto, Elena MartinoAbstract:Mutualistic and pathogenic plant-colonizing fungi use effector molecules to manipulate the host cell metabolism to allow plant tissue invasion. Some small secreted proteins (SSPs) have been identified as fungal effectors in both ectomycorrhizal and arbuscular mycorrhizal fungi, but it is currently unknown whether SSPs also play a role as effectors in other mycorrhizal associations. Ericoid mycorrhiza is a specific endomycorrhizal type that involves symbiotic fungi mostly belonging to the Leotiomycetes (Ascomycetes) and plants in the family Ericaceae. Genomic and RNASeq data from the ericoid mycorrhizal fungus Oidiodendron maius led to the identification of several symbiosis-upregulated genes encoding putative SSPs. OmSSP1, the most highly symbiosis up-regulated SSP, was found to share some features with fungal Hydrophobins, even though it lacks the Pfam hydrophobin domain. Sequence alignment with other Hydrophobins and hydrophobin-like fungal proteins placed OmSSP1 within Class I Hydrophobins. However, the predicted features of OmSSP1 may suggest a distinct type of hydrophobin-like proteins. The presence of a predicted signal peptide and a yeast-based signal sequence trap assay demonstrate that OmSSP1 is secreted. OmSSP1 null-mutants showed a reduced capacity to form ericoid mycorrhiza with Vaccinium myrtillus roots, suggesting a role as effectors in the ericoid mycorrhizal interaction.
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Phylogenetic, genomic organization and expression analysis of hydrophobin genes in the ectomycorrhizal basidiomycete Laccaria bicolor
Fungal genetics and biology : FG & B, 2012Co-Authors: Jonathan M. Plett, Julien Gibon, Annegret Kohler, Kecia Duffy, Patrik J. Hoegger, Rajesh Velagapudi, James Han, Ursula Kües, Igor V. Grigoriev, Francis MartinAbstract:Hydrophobins are morphogenetic, small secreted hydrophobic fungal proteins produced in response to changing development and environmental conditions. These proteins are important in the interaction between certain fungi and their hosts. In mutualistic ectomycorrhizal fungi several Hydrophobins form a subclass of mycorrhizal-induced small secreted proteins that are likely to be critical in the formation of the symbiotic interface with host root cells. In this study, two genomes of the ectomycorrhizal basidiomycete Laccaria bicolor strains S238N-H82 (from North America) and 81306 (from Europe) were surveyed to construct a comprehensive genome-wide inventory of Hydrophobins and to explore their characteristics and roles during host colonization. The S238N-H82 L. bicolor hydrophobin gene family is composed of 12 genes while the 81306 strain encodes nine Hydrophobins, all corresponding to class I Hydrophobins. The three extra hydrophobin genes encoded by the S238N-H82 genome likely arose via gene duplication and are bordered by transposon rich regions. Expression profiles of the hydrophobin genes of L. bicolor varied greatly depending on life stage (e.g. free living mycelium vs. root colonization) and on the host root environment. We conclude from this study that the complex diversity and range of expression profiles of the Laccaria hydrophobin multi-gene family have likely been a selective advantage for this mutualist in colonizing a wide range of host plants.
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Phylogenetic, genomic organization and expression analysis of hydrophobin genes in the ectomycorrhizal basidiomycete Laccaria bicolor
Fungal Genetics and Biology, 2012Co-Authors: Jonathan M. Plett, Julien Gibon, Annegret Kohler, Kecia Duffy, Patrik J. Hoegger, Rajesh Velagapudi, James Han, Igor V. Grigoriev, Ursula Kuees, Francis MartinAbstract:Hydrophobins are morphogenetic, small secreted hydrophobic fungal proteins produced in response to changing development and environmental conditions. These proteins are important in the interaction between certain fungi and their hosts. In mutualistic ectomycorrhizal fungi several Hydrophobins form a subclass of mycorrhizal-induced small secreted proteins that are likely to be critical in the formation of the symbiotic interface with host root cells. In this study, two genomes of the ectomycorrhizal basidiomycete Laccaria bicolor strains S238N-H82 (from North America) and 81306 (from Europe) were surveyed to construct a comprehensive genome-wide inventory of Hydrophobins and to explore their characteristics and roles during host colonization. The S238N-H82 L bicolor hydrophobin gene family is composed of 12 genes while the 81306 strain encodes nine Hydrophobins, all corresponding to class I Hydrophobins. The three extra hydrophobin genes encoded by the S238N-H82 genome likely arose via gene duplication and are bordered by transposon rich regions. Expression profiles of the hydrophobin genes oft. bicolor varied greatly depending on life stage (e.g. free living mycelium vs. root colonization) and on the host root environment. We conclude from this study that the complex diversity and range of expression profiles of the Laccaria hydrophobin multi-gene family have likely been a selective advantage for this mutualist in colonizing a wide range of host plants. (C) 2012 Elsevier Inc. All rights reserved.
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Cloning and expression analysis of a new hydrophobin cDNA from the ectomycorrhizal basidiomycete Pisolithus
Current Genetics, 2001Co-Authors: Sébastien Duplessis, Céline Sorin, Béatrice Palin, Catherine Voiblet, Francis Martin, Denis TaguAbstract:Hydrophobins are fungal cell wall proteins which play a crucial role in cell adhesion and aggregative processes. We have identified a new hydrophobin cDNA (hydPt-3) in the symbiotic mycelium of Pisolithus tinctorius (putative P. albus) during the formation of ectomycorrhizae around eucalypt roots. This sequence is highly divergent from two other previously identified Pisolithus symbiosis-regulated Hydrophobins, hydPt-1 and hydPt-2. Also, expression analyses demonstrated that hydPt-3 is up-regulated during the formation of ectomycorrhizae. In contrast to phytopathogenic fungi, changes in glucose or ammonium concentrations in the growth medium did not influence the accumulation of any Pisolithus hydrophobin mRNAs. This suggests that other factors act as regulators of hydrophobin gene expression in ectomycorrhizae.
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immunolocalization of hydrophobin hydpt 1 from the ectomycorrhizal basidiomycete pisolithus tinctorius during colonization of eucalyptus globulus roots
New Phytologist, 2001Co-Authors: Denis Tagu, O M H De Vries, Giovanni Piccoli, Ray Bellis, Paola Bonfante, Raffaella Balestrini, Vilberto Stocchi, Francis MartinAbstract:• The immunolocalization of one of the Hydrophobins of Pisolithustinctorius (HYDPt-1) is reported. Hydrophobin proteins play key roles in adhesion and aggregation of fungal hyphae, and it is already known that formation of ectomycorrhizas on eucalypt roots enhances the accumulation of hydrophobin mRNAs in the mycelium of Pisolithus tinctorius. • Purification of SDS-insoluble proteins from the mycelium of P. tinctorius showed the presence of a 13 kDa polypeptide with properties of class I hydrophobin. • Polyconal antibodies were raised against a recombinant HYDPt-1 polypeptide, and these were used for immunofluorescence-coupled transmission electron microscopy. • HYDPt-1 is a cell wall protein located at the surface of the hyphae with no preferential accumulation in the fungal cells of the different tissues of the ectomycorrhiza (i.e. extraradical hyphae, mantle or Hartig net).
Ann H. Kwan - One of the best experts on this subject based on the ideXlab platform.
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Fungal Hydrophobin Proteins Produce Self-Assembling Protein Films with Diverse Structure and Chemical Stability.
Nanomaterials (Basel Switzerland), 2014Co-Authors: Qin Ren, Ann H. Kwan, Chi L.l. Pham, Vanessa K. Morris, Margaret SundeAbstract:Hydrophobins are small proteins secreted by fungi and which spontaneously assemble into amphipathic layers at hydrophilic-hydrophobic interfaces. We have examined the self-assembly of the Class I Hydrophobins EAS∆15 and DewA, the Class II hydrophobin NC2 and an engineered chimeric hydrophobin. These Class I Hydrophobins form layers composed of laterally associated fibrils with an underlying amyloid structure. These two Class I Hydrophobins, despite showing significant conformational differences in solution, self-assemble to form fibrillar layers with very similar structures and require a hydrophilic-hydrophobic interface to trigger self-assembly. Addition of additives that influence surface tension can be used to manipulate the fine structure of the protein films. The Class II hydrophobin NC2 forms a mesh-like protein network and the engineered chimeric hydrophobin displays two multimeric forms, depending on assembly conditions. When formed on a graphite surface, the fibrillar EAS∆15 layers are resistant to alcohol, acid and basic washes. In contrast, the NC2 Class II monolayers are dissociated by alcohol treatment but are relatively stable towards acid and base washes. The engineered chimeric Class I/II hydrophobin shows increased stability towards alcohol and acid and base washes. Self-assembled hydrophobin films may have extensive applications in biotechnology where biocompatible; amphipathic coatings facilitate the functionalization of nanomaterials.
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Backbone and sidechain ^1H, ^13C and ^15N chemical shift assignments of the hydrophobin MPG1 from the rice blast fungus Magnaporthe oryzae
Biomolecular NMR Assignments, 2013Co-Authors: Anthony A. Rey, Antoine Hocher, Ann H. Kwan, Margaret SundeAbstract:Fungal Hydrophobins are secreted proteins that self-assemble at hydrophobic:hydrophilic interfaces. They are essential for a variety of processes in the fungal life cycle, including mediating interactions with surfaces and infection of hosts. The fungus Magnaporthe oryzae , the causative agent of rice blast, relies on the unique properties of Hydrophobins to infect cultivated rice as well as over 50 different grass species. The hydrophobin MPG1 is highly expressed during rice blast pathogenesis and has been implicated during host infection. Here we report the backbone and sidechain assignments for the class I hydrophobin MPG1 from the rice blast fungus Magnaporthe oryzae .
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analysis of the structure and conformational states of dewa gives insight into the assembly of the fungal Hydrophobins
Journal of Molecular Biology, 2013Co-Authors: Vanessa K. Morris, Ann H. Kwan, Margaret SundeAbstract:The hydrophobin DewA from the fungus Aspergillus nidulans is a highly surface-active protein that spontaneously self-assembles into amphipathic monolayers at hydrophobic:hydrophilic interfaces. These monolayers are composed of fibrils that are a form of functional amyloid. While there has been significant interest in the use of DewA for a variety of surface coatings and as an emulsifier in biotechnological applications, little is understood about the structure of the protein or the mechanism of self-assembly. We have solved the solution NMR structure of DewA. While the pattern of four disulfide bonds that is a defining feature of Hydrophobins is conserved, the arrangement and composition of secondary-structure elements in DewA are quite different to what has been observed in other hydrophobin structures. In addition, we demonstrate that DewA populates two conformations in solution, both of which are assembly competent. One conformer forms a dimer at high concentrations, but this dimer is off-pathway to fibril formation and may represent an assembly control mechanism. These data highlight the structural differences between fibril-forming Hydrophobins and those that form amorphous monolayers. This work will open up new opportunities for the engineering of Hydrophobins with novel biotechnological applications.
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Surface functionalization of carbon nanomaterials by self-assembling hydrophobin proteins.
Biopolymers, 2012Co-Authors: Wenrong Yang, Anthony A. Rey, Ann H. Kwan, Qin Ren, Vanessa K. Morris, Filip Braet, Margaret SundeAbstract:Class I fungal Hydrophobins are small surface-active proteins that self-assemble to form amphipathic monolayers composed of amyloid-like rodlets. The monolayers are extremely robust and can adsorb onto both hydrophobic and hydrophilic surfaces to reverse their wettability. This adherence is particularly strong for hydrophobic materials. In this report, we show that the class I Hydrophobins EAS and HYD3 can self-assemble to form a single-molecule thick coating on a range of nanomaterials, including single-walled carbon nanotubes (SWCNTs), graphene sheets, highly oriented pyrolytic graphite, and mica. Moreover, coating by class I hydrophobin results in a stable, dispersed preparation of SWCNTs in aqueous solutions. No cytotoxicity is detected when hydrophobin or hydrophobin-coated SWCNTs are incubated with Caco-2 cells in vitro. In addition, we are able to specifically introduce covalently linked chemical moieties to the hydrophilic side of the rodlet monolayer. Hence, class I Hydrophobins provide a simple and effective strategy for controlling the surfaces of a range of materials at a molecular level and exhibit strong potential for biomedical applications. © 2012 Wiley Periodicals, Inc.
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Backbone and sidechain 1H, 13C and 15N chemical shift assignments of the hydrophobin MPG1 from the rice blast fungus Magnaporthe oryzae
Biomolecular NMR assignments, 2012Co-Authors: Anthony A. Rey, Antoine Hocher, Ann H. Kwan, Margaret SundeAbstract:Fungal Hydrophobins are secreted proteins that self-assemble at hydrophobic:hydrophilic interfaces. They are essential for a variety of processes in the fungal life cycle, including mediating interactions with surfaces and infection of hosts. The fungus Magnaporthe oryzae, the causative agent of rice blast, relies on the unique properties of Hydrophobins to infect cultivated rice as well as over 50 different grass species. The hydrophobin MPG1 is highly expressed during rice blast pathogenesis and has been implicated during host infection. Here we report the backbone and sidechain assignments for the class I hydrophobin MPG1 from the rice blast fungus Magnaporthe oryzae.
Merja Penttilä - One of the best experts on this subject based on the ideXlab platform.
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direct identification of Hydrophobins and their processing in trichoderma using intact cell maldi tof ms
FEBS Journal, 2007Co-Authors: Torsten Neuhof, Merja Penttilä, Tiina Nakarisetala, Ralf Dieckmann, Irina S Druzhinina, Christian P Kubicek, Hans Von DohrenAbstract:Intact-cell MS (ICMS) was applied for the direct detection of Hydrophobins in various species and strains of Hypocrea/Trichoderma. In both mycelia and spores, dominating peaks were identified as Hydrophobins by detecting mass shifts of 8 Da of reduced and unreduced forms, the analysis of knockout mutants, and comparison with protein databases. Strain-specific processing was observed in the case of Hypocrea jecorina (anamorph Trichoderma reesei). An analysis of 32 strains comprising 29 different species of Trichoderma and Hypocrea showed hydrophobin patterns that were specific at both at the species and isolate (subspecies) levels. The method therefore permits rapid and direct detection of hydrophobin class II compositions and may also provide a means to identify Trichoderma (and other fungal) species and strains from microgram amounts of biomass without prior cultivation.
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Interaction and Comparison of a Class I Hydrophobin from Schizophyllum commune and Class II Hydrophobins from Trichoderma reesei
Biomacromolecules, 2006Co-Authors: Sanna Askolin, Markus Linder, Merja Penttilä, Karin Scholtmeijer, Marcel L. De Vocht, Maija Tenkanen, Han A. B. WöstenAbstract:Hydrophobins fulfill a wide spectrum of functions in fungal growth and development. These proteins self-assemble at hydrophilic-hydrophobic interfaces into amphipathic membranes. Hydrophobins are divided into two classes based on their hydropathy patterns and solubility. We show here that the properties of the class II Hydrophobins HFBI and HFBII of Trichoderma reesei differ from those of the class I hydrophobin SC3 of Schizophyllum commune. In contrast to SC3, self-assembly of HFBI and HFBII at the water-air interface was neither accompanied by a change in secondary structure nor by a change in ultrastructure. Moreover, maximal lowering of the water surface tension was obtained instantly or took several minutes in the case of HFBII and HFBI, respectively. In contrast, it took several hours in the case of SC3. Oil emulsions prepared with HFBI and SC3 were more stable than those of HFBII, and HFBI and SC3 also interacted more strongly with the hydrophobic Teflon surface making it wettable. Yet, the HFBI coating did not resist treatment with hot detergent, while that of SC3 remained unaffected. Interaction of all the Hydrophobins with Teflon was accompanied with a change in the circular dichroism spectra, indicating the formation of an alpha-helical structure. HFBI and HFBII did not affect self-assembly of the class I hydrophobin SC3 of S. commune and vice versa. However, precipitation of SC3 was reduced by the class II Hydrophobins, indicating interaction between the assemblies of both classes of Hydrophobins.
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Hydrophobins: the protein-amphiphiles of filamentous fungi.
FEMS microbiology reviews, 2005Co-Authors: Markus Linder, Tiina Nakari-setälä, Géza R. Szilvay, Merja PenttiläAbstract:Hydrophobins are surface active proteins produced by filamentous fungi. They have a role in fungal growth as structural components and in the interaction of fungi with their environment. They have, for example, been found to be important for aerial growth, and for the attachment of fungi to solid supports. Hydrophobins also render fungal structures, such as spores, hydrophobic. The biophysical properties of the isolated proteins are remarkable, such as strong adhesion, high surface activity and the formation of various self-assembled structures. The first high resolution three dimensional structure of a hydrophobin, HFBII from Trichoderma reesei, was recently solved. In this review, the properties of Hydrophobins are analyzed in light of these new data. Various application possibilities are also discussed.
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Fungal Hydrophobins as Predictors of the Gushing Activity of Malt
Journal of the Institute of Brewing, 2005Co-Authors: Tuija Sarlin, Markus Linder, Tiina Nakari-setälä, Merja Penttilä, Auli HaikaraAbstract:Fungal infection of barley and malt, particularly by strains of the genus Fusarium, is known to be a direct cause of beer gushing. We have shown previously that small fungal proteins, Hydrophobins, isolated from strains of the genera Fusarium, Nigrospora and Trichoderma act as gushing factors in beer. A hydrophobin concentration as low as 0.003 ppm was sufficient to induce gushing. The gushing-inducing abilities of the isolated Hydrophobins varied probably due to their structural differences. The Hydrophobins did not affect beer foam stability. A correlation was observed between the hydrophobin level analyzed by the hydrophobin ELISA developed and the gushing potential of malt. The risk of gushing was found to increase with hydrophobin concentrations above 250 μg/g malt. The levels of hydrophobin and the Fusarium mycotoxin deoxynivalenol (DON) in malts were not correlated which indicated that the formation of those two fungal metabolites may not be linked. Furthermore, we did not observe a correlation between the DON content and the gushing potential of the malt studied. Our observations suggest that the accuracy of predicting gushing could be improved by measuring the amount of the actual gushing factors, Hydrophobins, in barley or malt.
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Structural hierarchy in molecular films of two class II Hydrophobins
Biochemistry, 2003Co-Authors: Arja Paananen, Merja Penttilä, Elina Vuorimaa, Mika Torkkeli, Martti Kauranen, Olli Ikkala, Helge Lemmetyinen, Ritva Serimaa, Markus LinderAbstract:Hydrophobins are highly surface-active proteins that are specific to filamentous fungi. They function as coatings on various fungal structures, enable aerial growth of hyphae, and facilitate attachment to surfaces. Little is known about their structures and structure−function relationships. In this work we show highly organized surface layers of Hydrophobins, representing the most detailed structural study of hydrophobin films so far. Langmuir−Blodgett films of class II Hydrophobins HFBI and HFBII from Trichoderma reesei were prepared and analyzed by atomic force microscopy. The films showed highly ordered two-dimensional crystalline structures. By combining our recent results on small-angle X-ray scattering of hydrophobin solutions, we found that the unit cells in the films have dimensions similar to those of tetrameric aggregates found in solutions. Further analysis leads to a model in which the building blocks of the two-dimensional crystals are shape-persistent supramolecules consisting of four hydrop...
