Molecular Structure of Hydrophobins Studied with Site-Directed Mutagenesis and Vibrational Sum-Frequency Generation Spectroscopy

Meister, Konrad; Paananen, Arja; Speet, Bart; Lienemann, Michael; Bakker, Huib J.

doi:10.1021/acs.jpcb.7b08865

Cited by 12 publications

(22 citation statements)

References 31 publications

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“…To discern them, we further measured the amide III signals. It has been verified that the SFG amide III signals are capable of directly distinguishing α-helix, loop, coil, and turn-like structures in proteins at interfaces. ,− The spectral profiles of the loop and α-helical structures were well separated in the amide III spectra which appeared below and above 1260 cm –1 , respectively . In contrast, the amide III signals of coil and turn-like structures were very weak.…”

Section: Resultsmentioning

confidence: 87%

Misfolding of a Human Islet Amyloid Polypeptide at the Lipid Membrane Populates through β-Sheet Conformers without Involving α-Helical Intermediates

et al. 2019

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Amyloid formation has been implicated in many fatal diseases, but its mechanism remains to be clarified due to a lack of effective methods that can capture the transient intermediates. Here, we experimentally demonstrate that sum frequency generation vibrational spectroscopy can unambiguously discriminate the intermediates during amyloid formation at the lipid membrane in situ and in real time by combining the chiral amide I and achiral amide II and amide III spectral signals of the protein backbone. Such a combination can directly identify the formation of β-hairpin-like monomers and β-sheet oligomers and fibrils. A strong correlation between the amide II signals and the formation of β-sheet oligomers and fibrils was found. With this approach, the structural evolution of human islet amyloid polypeptides (hIAPP) at negative lipid bilayers was elucidated. It was firmly confirmed that hIAPP populates through β-sheet conformers without involving α-helical intermediates. The membrane-associated assembly of hIAPP proceeds by assembling with a β-hairpin-like monomer at the lipid bilayer surface, rather than by inserting the preassembled β-sheet oligomers in solution. This newly established protocol is ready to be utilized in revealing the mechanism of amyloid aggregation at the lipid membrane.

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Section: Resultsmentioning

confidence: 87%

Misfolding of a Human Islet Amyloid Polypeptide at the Lipid Membrane Populates through β-Sheet Conformers without Involving α-Helical Intermediates

et al. 2019

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“…Therefore, A k is linearly dependent on the number density N of adsorbed molecules and the orientational average of the molecules’ hyperpolarizability β k . Because the second-order χ (2) and third-order χ (3) contributions are resonantly enhanced when the frequency ω IR is tuned over vibrational modes of interfacial molecules, these equations show how SFG spectroscopy is molecule specific and thus provides qualitative information on the composition 43 − 46 and the charging state 13 , 47 , 48 of proteins at interfaces.…”

Section: Resultsmentioning

confidence: 98%

Charge-Controlled Surface Properties of Native and Fluorophore-Labeled Bovine Serum Albumin at the Air–Water Interface

2018

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Proteins at interfaces are important for protein formulations and in soft materials such as foam. Here, interfacial stability and physicochemical properties are key elements, which drive macroscopic foam properties through structure–property relations. Native and fluorescein isothiocyanate-labeled bovine serum albumin (BSA) were used to modify air–water interfaces as a function of pH. Characterizations were performed with tensiometry and sum-frequency generation (SFG). SFG spectra of O–H stretching vibrations reveal a phase reversal and a pronounced minimum in O–H intensity at pH values of 5.3 and 4.7 for native and labeled BSA, respectively. This minimum is attributed to the interfacial isoelectric point (IEP) and is accompanied by a minimum in surface tension and negligible ζ-potentials in the bulk. Interfacial proteins at pH values close to the IEP can promote macroscopic foam stability and are predominately located in the lamellae between individual gas bubbles as evidenced by confocal fluorescence microscopy. Different from the classical stabilization mechanisms, for example, via the electrostatic disjoining pressure, we propose that the presence of more close-packed BSA, because of negligible net charges, inside the foam lamellae is more effective in reducing foam drainage as compared to a situation with strong repulsive electrostatic interactions.

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“…1e), respectively. It was shown previously that these mutations affect the interactions of the proteins laterally within an interfacial layer as well as with the water phase but do not alter the folding state of the proteins 31,32 .…”

Section: Proteinsmentioning

confidence: 95%

“…5a). Although the mutations involving the charged residues alter the mutual protein interactions as well as the interaction with the surrounding water phase 31,32 , stable bilayers were able to be produced from all variants. Like in the case of wild type HFBI, the determined bilayer tension values Γ (see fig.…”

Section: Van Der Waals Is the Dominant Contribution Of The Adhesion Ementioning

confidence: 99%

Adhesion Properties of Freestanding Hydrophobin Bilayers

et al. 2018

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Hydrophobins are a family of small-sized proteins featuring a distinct hydrophobic patch on the protein's surface, rendering them amphiphilic. This particularity allows hydrophobins to self-assemble into monolayers at any hydrophilic/hydrophobic interface. Moreover, stable pure protein bilayers can be created from two interfacial hydrophobin monolayers by contacting either their hydrophobic or their hydrophilic sides. In this study, this is achieved via a microfluidic approach, in which also the bilayers' adhesion energy can be determined. This enables us to study the origin of the adhesion of hydrophobic and hydrophilic core bilayers made from the class II hydrophobins HFBI and HFBII. Using different fluid media in this setup and introducing genetically modified variants of the HFBI molecule, the different force contributions to the adhesion of the bilayer sheets are studied. It was found that in the hydrophilic contact situation, the adhesive interaction was higher than that in the hydrophobic contact situation and could be even enhanced by reducing the contributions of electrostatic interactions. This effect indicates that the van der Waals interaction is the dominant contribution that explains the stability of the observed bilayers.

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Molecular Structure of Hydrophobins Studied with Site-Directed Mutagenesis and Vibrational Sum-Frequency Generation Spectroscopy

Cited by 12 publications

References 31 publications

Misfolding of a Human Islet Amyloid Polypeptide at the Lipid Membrane Populates through β-Sheet Conformers without Involving α-Helical Intermediates

Misfolding of a Human Islet Amyloid Polypeptide at the Lipid Membrane Populates through β-Sheet Conformers without Involving α-Helical Intermediates

Charge-Controlled Surface Properties of Native and Fluorophore-Labeled Bovine Serum Albumin at the Air–Water Interface

Adhesion Properties of Freestanding Hydrophobin Bilayers

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