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Surface-water Interface Induces Conformational Changes Critical for Protein Adsorption: Implications for Monolayer Formation of EAS Hydrophobin.

Ley K, Christofferson A, Penna M, Winkler D, Maclaughlin S, Yarovsky I - Front Mol Biosci (2015)

Bottom Line: The class I hydrophobin EAS is part of a family of small, amphiphilic fungal proteins best known for their ability to self-assemble into stable monolayers that modify the hydrophobicity of a surface to facilitate further microbial growth.Specific and water mediated interactions with the surface were also analyzed.We have identified two possible binding motifs, one which allows unfolding of the Cys7-Cys8 loop due to the surfactant-like behavior of the Cys3-Cys4 loop, and another which has limited unfolding due to the Cys3-Cys4 loop remaining disordered in solution.

View Article: PubMed Central - PubMed

Affiliation: Health Innovations Research Institute and School of Aerospace, Mechanical and Manufacturing Engineering, RMIT University Melbourne, VIC, Australia.

ABSTRACT
The class I hydrophobin EAS is part of a family of small, amphiphilic fungal proteins best known for their ability to self-assemble into stable monolayers that modify the hydrophobicity of a surface to facilitate further microbial growth. These proteins have attracted increasing attention for industrial and biomedical applications, with the aim of designing surfaces that have the potential to maintain their clean state by resisting non-specific protein binding. To gain a better understanding of this process, we have employed all-atom molecular dynamics to study initial stages of the spontaneous adsorption of monomeric EAS hydrophobin on fully hydroxylated silica, a commonly used industrial and biomedical substrate. Particular interest has been paid to the Cys3-Cys4 loop, which has been shown to exhibit disruptive behavior in solution, and the Cys7-Cys8 loop, which is believed to be involved in the aggregation of EAS hydrophobin at interfaces. Specific and water mediated interactions with the surface were also analyzed. We have identified two possible binding motifs, one which allows unfolding of the Cys7-Cys8 loop due to the surfactant-like behavior of the Cys3-Cys4 loop, and another which has limited unfolding due to the Cys3-Cys4 loop remaining disordered in solution. We have also identified intermittent interactions with water which mediate the protein adsorption to the surface, as well as longer lasting interactions which control the diffusion of water around the adsorption site. These results have shown that EAS behaves in a similar way at the air-water and surface-water interfaces, and have also highlighted the need for hydrophilic ligand functionalization of the silica surface in order to prevent the adsorption of EAS hydrophobin.

No MeSH data available.


Mean squared displacement (MSD) plots of: (A) water molecules at the, air-water, bulk water and surface-water interface; both with and without the presence of EAS hydrophobin. (B) lateral MSD of the protein at the air-water interface, in bulk solution, and in both binding motifs at the surface-water interface.
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Figure 8: Mean squared displacement (MSD) plots of: (A) water molecules at the, air-water, bulk water and surface-water interface; both with and without the presence of EAS hydrophobin. (B) lateral MSD of the protein at the air-water interface, in bulk solution, and in both binding motifs at the surface-water interface.

Mentions: The three-dimensional mean squared displacement (MSD) of water molecules at the surface-water interface and air-water interface with and without the presence of EAS have been compared to that of bulk water (Figure 8A). The curves are generated over a short-time domain (10 ps), with the gradient from a line of best fit plot proportional to the diffusion coefficient for the water molecules in the respective zones (Yiapanis et al., 2013). We observe a diffusion coefficient of 4.3 × 10−5 cm2/s for bulk water (Figure 8A), which is slightly higher than the reported 4.0 × 10−5 cm2/s diffusion coefficient for TIP3P with Ewald summation (Price and Brooks, 2004). As expected, the water at the surface-water interface (2.35 × 10−5 cm2/s) is significantly slower than bulk, due to stabilizing interactions with the silica surface. This is again reduced further when the protein is present (2.10 × 10−5 cm2/s), which shows that the protein does in fact trap water in the adsorption region, and limit the diffusion of water through aforementioned long polar and charged side-chain residues that are 5–6 Å from the surface, such as aspartic acid and serine. Conversely, at the air-water interface the diffusion is a factor of 10 faster without the protein, (1.15 × 10−4 cm2/s) slowing significantly in the presence of the protein (6.75 × 10−5 cm2/s). This behavior is in turn replicated for the mobility of the protein itself, where the MSD of the protein at the two interfaces can be seen in Figure 8B. At the surface-water interface the protein is practically immobile on the surface, with very little movement occurring once the protein is adsorbed. Comparatively at the air-water interface the protein moves at the interface freely, suggesting that there may be different mechanisms involved for hydrophobin monolayer formation at the air-water and surface-water interface, due to the vastly different surface hydropathicity and mobility of the protein. It may provide further support to the theory that the Cys3-Cys4 loop has surfactant-like behavior at the air-water interface (De Simone et al., 2012), especially considering the significant effects it has on the water diffusion coefficient.


Surface-water Interface Induces Conformational Changes Critical for Protein Adsorption: Implications for Monolayer Formation of EAS Hydrophobin.

Ley K, Christofferson A, Penna M, Winkler D, Maclaughlin S, Yarovsky I - Front Mol Biosci (2015)

Mean squared displacement (MSD) plots of: (A) water molecules at the, air-water, bulk water and surface-water interface; both with and without the presence of EAS hydrophobin. (B) lateral MSD of the protein at the air-water interface, in bulk solution, and in both binding motifs at the surface-water interface.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4644811&req=5

Figure 8: Mean squared displacement (MSD) plots of: (A) water molecules at the, air-water, bulk water and surface-water interface; both with and without the presence of EAS hydrophobin. (B) lateral MSD of the protein at the air-water interface, in bulk solution, and in both binding motifs at the surface-water interface.
Mentions: The three-dimensional mean squared displacement (MSD) of water molecules at the surface-water interface and air-water interface with and without the presence of EAS have been compared to that of bulk water (Figure 8A). The curves are generated over a short-time domain (10 ps), with the gradient from a line of best fit plot proportional to the diffusion coefficient for the water molecules in the respective zones (Yiapanis et al., 2013). We observe a diffusion coefficient of 4.3 × 10−5 cm2/s for bulk water (Figure 8A), which is slightly higher than the reported 4.0 × 10−5 cm2/s diffusion coefficient for TIP3P with Ewald summation (Price and Brooks, 2004). As expected, the water at the surface-water interface (2.35 × 10−5 cm2/s) is significantly slower than bulk, due to stabilizing interactions with the silica surface. This is again reduced further when the protein is present (2.10 × 10−5 cm2/s), which shows that the protein does in fact trap water in the adsorption region, and limit the diffusion of water through aforementioned long polar and charged side-chain residues that are 5–6 Å from the surface, such as aspartic acid and serine. Conversely, at the air-water interface the diffusion is a factor of 10 faster without the protein, (1.15 × 10−4 cm2/s) slowing significantly in the presence of the protein (6.75 × 10−5 cm2/s). This behavior is in turn replicated for the mobility of the protein itself, where the MSD of the protein at the two interfaces can be seen in Figure 8B. At the surface-water interface the protein is practically immobile on the surface, with very little movement occurring once the protein is adsorbed. Comparatively at the air-water interface the protein moves at the interface freely, suggesting that there may be different mechanisms involved for hydrophobin monolayer formation at the air-water and surface-water interface, due to the vastly different surface hydropathicity and mobility of the protein. It may provide further support to the theory that the Cys3-Cys4 loop has surfactant-like behavior at the air-water interface (De Simone et al., 2012), especially considering the significant effects it has on the water diffusion coefficient.

Bottom Line: The class I hydrophobin EAS is part of a family of small, amphiphilic fungal proteins best known for their ability to self-assemble into stable monolayers that modify the hydrophobicity of a surface to facilitate further microbial growth.Specific and water mediated interactions with the surface were also analyzed.We have identified two possible binding motifs, one which allows unfolding of the Cys7-Cys8 loop due to the surfactant-like behavior of the Cys3-Cys4 loop, and another which has limited unfolding due to the Cys3-Cys4 loop remaining disordered in solution.

View Article: PubMed Central - PubMed

Affiliation: Health Innovations Research Institute and School of Aerospace, Mechanical and Manufacturing Engineering, RMIT University Melbourne, VIC, Australia.

ABSTRACT
The class I hydrophobin EAS is part of a family of small, amphiphilic fungal proteins best known for their ability to self-assemble into stable monolayers that modify the hydrophobicity of a surface to facilitate further microbial growth. These proteins have attracted increasing attention for industrial and biomedical applications, with the aim of designing surfaces that have the potential to maintain their clean state by resisting non-specific protein binding. To gain a better understanding of this process, we have employed all-atom molecular dynamics to study initial stages of the spontaneous adsorption of monomeric EAS hydrophobin on fully hydroxylated silica, a commonly used industrial and biomedical substrate. Particular interest has been paid to the Cys3-Cys4 loop, which has been shown to exhibit disruptive behavior in solution, and the Cys7-Cys8 loop, which is believed to be involved in the aggregation of EAS hydrophobin at interfaces. Specific and water mediated interactions with the surface were also analyzed. We have identified two possible binding motifs, one which allows unfolding of the Cys7-Cys8 loop due to the surfactant-like behavior of the Cys3-Cys4 loop, and another which has limited unfolding due to the Cys3-Cys4 loop remaining disordered in solution. We have also identified intermittent interactions with water which mediate the protein adsorption to the surface, as well as longer lasting interactions which control the diffusion of water around the adsorption site. These results have shown that EAS behaves in a similar way at the air-water and surface-water interfaces, and have also highlighted the need for hydrophilic ligand functionalization of the silica surface in order to prevent the adsorption of EAS hydrophobin.

No MeSH data available.