pH-induced insertion of pHLIP into a lipid bilayer: In-situ SEIRAS characterization of a folding intermediate at neutral pH

Ataka, Kenichi; Drauschke, Janina; Stulberg, Valentina; Koksch, Beate; Heberle, Joachim

doi:10.1016/j.bbamem.2022.183873

Cited by 18 publications

(11 citation statements)

References 34 publications

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“…The lipid‐modified SEIRA chip was submitted to a PBS buffer and the peptides were subsequently added. The sample SEIRA spectra were simultaneously measured to observe adsorption/folding process of the polypeptides on the solid‐ supported lipid bilayer [27] . For these studies, the X13 K peptides were selected, since here, both all degrees of fluorination were present and the structural diversity within the series was the most pronounced.…”

Section: Resultsmentioning

confidence: 99%

“…Since conventional FTIR spectroscopy is not sufficiently sensitive to detect the peptide oligomers at the given concentration, we employed Surface-Enhanced Absorption Spectroscopy (SEIRAS). [27] For this purpose, large unilamellar vesicles (LUV) of POPC/POPG (1 : 1) were casted over the SEIRA active gold film surface to form a solid-supported lipid bilayer. The lipidmodified SEIRA chip was submitted to a PBS buffer and the peptides were subsequently added.…”

Section: Surface Enhance Infrared Absorption Spectroscopy (Seiras)mentioning

confidence: 99%

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Introducing Aliphatic Fluoropeptides: Perspectives on Folding Properties, Membrane Partition and Proteolytic Stability

Hohmann

Chowdhary

Ataka

et al. 2023

Chemistry A European J

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A de novo designed class of peptide-based fluoropolymers composed of fluorinated aliphatic amino acids as main components is reported. Structural characterization provided insights into fluorine-induced alterations on βstrand to α-helix transition upon an increase in SDS content and revealed the unique formation of PPII structures for trifluorinated fluoropeptides. A combination of circular dichroism, fluorescence-based leaking assays and surface enhanced infrared absorption spectroscopy served to examine the insertion and folding processes into unilamellar vesicles. While partitioning into lipid bilayers, the degree of fluorination conducts a decrease in α-helical content. Furthermore, this study comprises a report on the proteolytic stability of peptides exclusively built up by fluorinated amino acids and proved all sequences to be enzymatically degradable despite the degree of fluorination. Herein presented fluoropeptides as well as the distinctive properties of these artificial and polyfluorinated foldamers with enzyme-degradable features will play a crucial role in the future development of fluorinated peptide-based biomaterials.

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Section: Resultsmentioning

confidence: 99%

Section: Surface Enhance Infrared Absorption Spectroscopy (Seiras)mentioning

confidence: 99%

Introducing Aliphatic Fluoropeptides: Perspectives on Folding Properties, Membrane Partition and Proteolytic Stability

Hohmann

Chowdhary

Ataka

et al. 2023

Chemistry A European J

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“…solid‐supported bilayer lipid membrane ; Abbildung 1) als Minimalmodellsystem der Virusmembran eingebettet [22,23] . SEIRA‐Spektroskopie ermöglicht die Untersuchung von Protonierungsereignissen [24,25] sowie die Quantifizierung weitreichender helikaler Reorientierungen [26,27] . Dies wird durch einen nanostrukturierten Goldfilm erreicht, der eine plasmonische Infrarot‐(IR)‐Signalverstärkung der rekonstituierten Proteine in einer einzelnen ssBLM um einen Faktor von 10 2 bewirkt [28,29] .…”

Section: Figureunclassified

In situ‐spektroskopische Detektion der weitreichenden Reorientierung von Transmembranhelices während der Öffnung des Influenza A M2‐Kanals**

Paschke,

Mohr,

Lange

et al. 2023

Angewandte Chemie

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Viroporins are small ion channels in membranes of enveloped viruses that play key roles during viral life cycles. To use viroporins as drug targets against viral infection requires in‐depth mechanistic understanding and, with that, methods that enable investigations under in situ conditions. Here, we apply surface‐enhanced infrared absorption (SEIRA) spectroscopy to Influenza A M2 reconstituted within a solid‐supported membrane, to shed light on the mechanics of its viroporin function. M2 is a paradigm of pH‐activated proton channels and controls the proton flux into the viral interior during viral infection. We use SEIRA to track the large‐scale reorientation of M2’s transmembrane α‐helices in situ during pH‐activated channel opening. We quantify this event as a helical tilt from 26° to 40° by correlating the experimental results with solid‐state nuclear magnetic resonance‐informed computational spectroscopy. This mechanical motion is impeded upon addition of the inhibitor rimantadine, giving a direct spectroscopic marker to test antiviral activity. The presented approach provides a spectroscopic tool to quantify large‐scale structural changes and to track the function and inhibition of the growing number of viroporins from pathogenic viruses in future studies.

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“…7 At more basic conditions, pHLIP peptides typically unfold to a random coil conformation, either in solution (pH > 8.0) (State I) or adsorbed to the membrane surface (pH 7.0 to 8.0) (State II). 2 Although the folding process has been challenging to describe, as shown in recent work regarding the insertion transition states, 8 the characteristic pH dependency relates to key titrating residues identified as regulators of this insertion process, such as Asp-14 in the wt peptide, and their local interactions with the surrounding environment. Indeed, during the design process of several variant sequences, an aspartate residue found in a key location was essential.…”

Section: Introductionmentioning

confidence: 99%

Increasing the Realism of in Silico pHLIP Peptide Models with a Novel pH Gradient CpHMD Method

Silva

Vila-Viçosa

Machuqueiro

2022

J. Chem. Theory Comput.

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The pH-low insertion peptides (pHLIP) are pH-dependent membrane inserting peptides, whose function depends on the cell microenvironment acidity. Several peptide variants have been designed to improve upon the wt-sequence, particularly the state transition kinetics and the selectivity for tumor pH. The variant 3 (Var3) peptide is a 27 residue long peptide, with a key titrating residue (Asp-13) that, despite showing a modest performance in liposomes (pK ins ∼ 5.0), excelled in tumor cell experiments. To help rationalize these results, we focused on the pH gradient in the cell membrane, which is one of the crucial properties that are not present in liposomes. We extended our CpHMD-L method and its pH replica-exchange (pHRE) implementation to include a pH gradient and mimic the pHLIP-membrane microenvironment in a cell where the internal pH is fixed (pH 7.2) and the external pH is allowed to change. We showed that, by properly modeling the pH-gradient, we can correctly predict the experimentally observed loss and gain of performance in tumor cells experiments by the wt and Var3 sequences, respectively. In sum, the pH gradient implementation allowed for more accurate and realistic pK a estimations and was a pivotal step in bridging the in silico data and the in vivo cell experiments.

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pH-induced insertion of pHLIP into a lipid bilayer: In-situ SEIRAS characterization of a folding intermediate at neutral pH

Cited by 18 publications

References 34 publications

Introducing Aliphatic Fluoropeptides: Perspectives on Folding Properties, Membrane Partition and Proteolytic Stability

Introducing Aliphatic Fluoropeptides: Perspectives on Folding Properties, Membrane Partition and Proteolytic Stability

In situ‐spektroskopische Detektion der weitreichenden Reorientierung von Transmembranhelices während der Öffnung des Influenza A M2‐Kanals**

Increasing the Realism of in Silico pHLIP Peptide Models with a Novel pH Gradient CpHMD Method

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