Predicting the self-assembly film structure of class II hydrophobin NC2 and estimating its structural characteristics.

Hydrophobins are fungal proteins that can mediate water surface tension by forming amphiphilic self-assembly structures in hydrophobic-hydrophilic interfaces. Hydrophobins are known to self-assemble into two forms depending on their class: class I hydrophobins aggregate into a functional amyloid rodlet, while class II hydrophobins aggregate into a regularly patterned monolayer. Owing to its unique properties, hydrophobin has been considered as a biocompatible nanomaterial for various applications and there have been several attempts to engineer hydrophobins to enhance their function. Recently, a chimeric hydrophobin named NChi2 was found to be able to self-assemble into both rodlet and monolayer forms depending on the incubating environment. Although this remarkable feature suggests that NChi2 can function as a versatile bionanomaterial for various applications, only little information about the protein, such as its assembly structure or its characteristics, is provided. To investigate the extraordinary behavior of NChi2, it seems to be a prerequisite to first understand the characteristics of its parent hydrophobins, namely class I EAS and class II NC2. Here, we conducted a preliminary study on predicting the self-assembly structure of class II hydrophobin NC2 and estimating its structural characteristics by employing several computational methods. From the results, we found that NC2 shows stronger surface activity than HFBII, while its assembly structure is weaker than that of HFBII. We hope that this research serves as a foundation to further investigate the structural characteristics of a unique hydrophobin NChi2 in future studies.

Investigation of the role hydrophobin monomer loops using hybrid models via molecular dynamics simulation.

Hydrophobins are small amphiphilic fungal proteins that are highly surface-active and are used in various industrial applications such as dispersion, immobilization, and antifouling. At hydrophobic-hydrophilic interfaces, hydrophobins tend to self-assemble as rodlets or monolayers, depending on whether they are class I or II. Several studies have determined the three-dimensional structure and investigated the self-assembly formation mechanism of the class I EAS from Neurospora crassa and the class II HFBII from Trichoderma reesei. Although some studies have examined the performance of chimeric hydrophobins, they have not been investigated at the atomic scale. Here, we designed chimeric hydrophobins by grafting the L1 loop of Vmh2 and the L3 loop of EAS onto the class II hydrophobin HFBII by homology modeling and performed vacuum-water interface molecular simulations to determine their structural behaviors. We found that the chimeric hydrophobin grafted with the L3 of EAS became unstable under standard conditions, whereas that grafted with the L1 of Vmh2 became unstable in the presence of calcium ions. Moreover, when both the EAS L3 and Vmh2 L1 were grafted together, the structure became disordered and lost its amphiphilic characteristics in standard conditions. In the presence of calcium, however, its structural stability was restored. However, an additional external perturbation is required to trigger the conformational transition. Although our chimeric hydrophobin models were designed through homology modeling, our results provide detailed information regarding hydrophobin self-assembly and their surface-interactive behavior that may serve as a template for designing hydrophobins for future industrial applications.

Parenteral nanosuspensions: a brief review from solubility enhancement to more novel and specific applications.

Advancements in in silico techniques of lead molecule selection have resulted in the failure of around 70% of new chemical entities (NCEs). Some of these molecules are getting rejected at final developmental stage resulting in wastage of money and resources. Unfavourable physicochemical properties affect ADME profile of any efficacious and potent molecule, which may ultimately lead to killing of NCE at final stage. Numerous techniques are being explored including nanocrystals for solubility enhancement purposes. Nanocrystals are the most successful and the ones which had a shorter gap between invention and subsequent commercialization of the first marketed product. Several nanocrystal-based products are commercially available and there is a paradigm shift in using approach from simply being solubility enhancement technique to more novel and specific applications. Some other aspects in relation to parenteral nanosuspensions are concentrations of surfactant to be used, scalability and in vivo fate. At present, there exists a wide gap due to poor understanding of these critical factors, which we have tried to address in this review. This review will focus on parenteral nanosuspensions, covering varied aspects especially stabilizers used, GRAS (Generally Recognized as Safe) status of stabilizers, scalability challenges, issues of physical and chemical stability, solidification techniques to combat stability problems and in vivo fate.

The dynamics of multimer formation of the amphiphilic hydrophobin protein HFBII.

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 Förster 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.