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Recruitment of class I hydrophobins to the air:water interface initiates a multi-step process of functional amyloid formation.

ABSTRACT: Class I fungal hydrophobins form amphipathic monolayers composed of amyloid rodlets. This is a remarkable case of functional amyloid formation in that a hydrophobic:hydrophilic interface is required to trigger the self-assembly of the proteins. The mechanism of rodlet formation and the role of the interface in this process have not been well understood. Here, we have studied the effect of a range of additives, including ionic liquids, alcohols, and detergents, on rodlet formation by two class I hydrophobins, EAS and DewA. Although the conformation of the hydrophobins in these different solutions is not altered, we observe that the rate of rodlet formation is slowed as the surface tension of the solution is decreased, regardless of the nature of the additive. These results suggest that interface properties are of critical importance for the recruitment, alignment, and structural rearrangement of the amphipathic hydrophobin monomers. This work gives insight into the forces that drive macromolecular assembly of this unique family of proteins and allows us to propose a three-stage model for the interface-driven formation of rodlets.

SUBMITTER: Morris VK

PROVIDER: S-EPMC3091204 | biostudies-literature | 2011 May

REPOSITORIES: biostudies-literature

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Recruitment of class I hydrophobins to the air:water interface initiates a multi-step process of functional amyloid formation.

Morris Vanessa K VK Ren Qin Q Macindoe Ingrid I Kwan Ann H AH Byrne Nolene N Sunde Margaret M

The Journal of biological chemistry 20110318 18

Class I fungal hydrophobins form amphipathic monolayers composed of amyloid rodlets. This is a remarkable case of functional amyloid formation in that a hydrophobic:hydrophilic interface is required to trigger the self-assembly of the proteins. The mechanism of rodlet formation and the role of the interface in this process have not been well understood. Here, we have studied the effect of a range of additives, including ionic liquids, alcohols, and detergents, on rodlet formation by two class I ...[more]

PMID: 21454575

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Project description:Hydrophobins (HPs) are small secreted fungal proteins possibly involved in several processes such as formation of fungal aerial structures, attachment to hydrophobic surfaces, interaction with the environment and protection against the host defense system. The genome of the necrotrophic plant pathogen Fusarium graminearum contains five genes encoding for HPs (FgHyd1-5). Single and triple FgHyd mutants were produced and characterized. A reduced growth was observed when the ΔFghyd2 and the three triple mutants including the deletion of FgHyd2 were grown in complete or minimal medium. Surprisingly, the growth of these mutants was similar to wild-type when grown under ionic, osmotic or oxidative stress conditions. All the mutant strains confirmed the ability to develop conidia and perithecia, suggesting that the FgHyds are not involved in normal development of asexual and sexual structures. A reduction in the ability of hyphae to penetrate through the water-air interface was observed for the single mutants ΔFghyd2 and ΔFghyd3 as well as for the triple mutants including the deletion of FgHyd2 and FgHyd3. Besides, ΔFghyd3 and the triple mutant ΔFghyd234 were also affected in the attachment to hydrophobic surface. Indeed, wheat infection experiments showed a reduction of symptomatic spikelets for ΔFghyd2 and ΔFghyd3 and the triple mutants only when spray inoculation was performed. This result could be ascribed to the affected ability of mutants deleted of FgHyd2 and FgHyd3 to penetrate through the water-air interface and to attach to hydrophobic surfaces such as the spike tissue. This hypothesis is strengthened by a histological analysis, performed by fluorescence microscopy, showing no defects in the morphology of infection structures produced by mutant strains. Interestingly, triple hydrophobin mutants were significantly more inhibited than wild-type by the treatment with a systemic triazole fungicide, while no defects at the cell wall level were observed.

Project description:At the air/buffer solution interface the kinetics of adsorption of amyloid beta peptide, Abeta(1-42), whose bulk concentration (submicromolar) is more than 2 orders of magnitude lower than that typically used in other in vitro aggregation studies, has been studied using a Langmuir-Blodgett trough. The pressure-time curves exhibit a lag phase, wherein the surface pressure essentially remains at zero, and a rising phase, corresponding to the Abeta adsorption at the interface. The duration of the lag phase was found to be highly dependent on both the Abeta bulk concentration and the solution temperature. A large activation energy (62.2 +/- 4.1 KJ/mol) was determined and the apparent adsorption rate constant was found to be linearly dependent on the Abeta bulk concentration. Attenuated total reflection-IR spectra of the adsorbed Abeta transferred to a solid substrate and circular dichroism measurements of Abeta in the solution layer near the interface reveal that the natively unstructured Abeta in the bulk undergo a conformation change (folding) to mainly the alpha-helical structure. The results suggest that, prior to the adsorption step, an equilibrium between Abeta conformations is established within the subsurface. The kinetic equation derived from this model confirms that the overall Abeta adsorption is kinetically controlled and the apparent rate constant is proportional to the Abeta bulk concentration. This model also indicates that interfaces such as cell membranes and lipid bilayers may facilitate Abeta aggregation/ fibrillation by providing a thin hydrophobic layer adjacent to the interface for the initial A/beta conformation change (misfolding) and accumulation. Such a preconcentration effect offers a plausible explanation of the fact that Abeta fibrillation occurs in vivo at nanomolar concentrations. Another important biological implication from our work is that Abeta misfolding may occur before its adsorption onto a cell membrane. This general kinetic model should also find applications in adsorption studies of other types of biomolecules whose overall kinetics exhibits a lag phase that is dependent on the bulk concentration of the adsorbate.

Dataset Information

Recruitment of class I hydrophobins to the air:water interface initiates a multi-step process of functional amyloid formation.

Publications

Recruitment of class I hydrophobins to the air:water interface initiates a multi-step process of functional amyloid formation.

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