Protein Nanopore Membranes Prepared by a Simple Langmuir–Schaefer Approach

Schwieters, Magnus S.; Mathieu-Gaedke, Maria; Westphal, Michael I.; Dalpke, Raphael; Dirksen, Maxim; Qi, Daizong; Grull, Marco; Bick, Thomas; Tassler, Stephanie; Sauer, Daniel F.; Bonn, Mischa; Wendler, Petra; Hellweg, Thomas; Beyer, André; Schwaneberg, Ulrich; Glebe, Ulrich; Böker, Alexander

doi:10.1002/smll.202102975

Cited by 3 publications

(3 citation statements)

References 61 publications

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“…Such assemblies at the oil–water interface using transmembrane proteins are of interest to generating protocells because the channel of FhuA might act as a gateway in and out of the protocell, thereby mimicking real living cells. Moreover, we demonstrated that polymeric protein membranes can be formed by directly cross-linking FhuA with glutaraldehyde …”

Section: Fhua In Nanobiotechnology and Materials Designmentioning

confidence: 99%

“…Another exciting application of FhuA is the possibility of incorporating FhuA into polymers, ,,,, as already discussed above. A catalytically active membrane can be constructed by incorporating a FhuA-based artificial metalloenzyme into a polymer matrix .…”

Section: Fhua As a Protein Scaffold For Metal Catalystsmentioning

confidence: 99%

“…Moreover, we demonstrated that polymeric protein membranes can be formed by directly cross-linking FhuA with glutaraldehyde. 25…”

Section: Fhua As a Building Block In Copolymerizationsmentioning

confidence: 99%

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FhuA: From Iron-Transporting Transmembrane Protein to Versatile Scaffolds through Protein Engineering

et al. 2023

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Metrics & MoreArticle Recommendations CONSPECTUS: Protein engineering has emerged as a powerful methodology to tailor the properties of proteins. It empowers the design of biohybrid catalysts and materials, thereby enabling the convergence of materials science, chemistry, and medicine. The choice of a protein scaffold is an important factor for performance and potential applications. In the past two decades, we utilized the ferric hydroxamate uptake protein FhuA. FhuA is, from our point of view, a versatile scaffold due to its comparably large cavity and robustness toward temperature as well as organic cosolvents. FhuA is a natural iron transporter located in the outer membrane of Escherichia coli (E. coli). Wild-type FhuA consists of 714 amino acids and has a β-barrel structure composed of 22 antiparallel β-sheets, closed by an internal globular "cork" domain (amino acids 1−160). FhuA is robust in a broad pH range and toward organic cosolvents; therefore, we envisioned FhuA to be a suitable platform for various applications in (i) biocatalysis, (ii) materials science, and (iii) the construction of artificial metalloenzymes.(i) Applications in biocatalysis were achieved by removing the globular cork domain (FhuA_Δ1−160), thereby creating a large pore for the passive transport of otherwise difficult-to-import molecules through diffusion. Introducing this FhuA variant into the outer membrane of E. coli facilitates the uptake of substrates for downstream biocatalytic conversion. Furthermore, removing the globular "cork" domain without structural collapse of the ß-barrel protein allowed the use of FhuA as a membrane filter, exhibiting a preference for D-arginine over L-arginine.(ii) FhuA is a transmembrane protein, which makes it attractive to be used for applications in non-natural polymeric membranes. Inserting FhuA into polymer vesicles yielded so-called synthosomes (i.e., catalytic synthetic vesicles in which the transmembrane protein acted as a switchable gate or filter). Our work in this direction enables polymersomes to be used in biocatalysis, DNA recovery, and the controlled (triggered) release of molecules. Furthermore, FhuA can be used as a building block to create protein− polymer conjugates to generate membranes.(iii) Artificial metalloenzymes (ArMs) are formed by incorporating a non-native metal ion or metal complex into a protein. This combines the best of two worlds: the vast reaction and substrate scope of chemocatalysis and the selectivity and evolvability of enzymes. With its large inner diameter, FhuA can harbor (bulky) metal catalysts. Among others, we covalently attached a Grubbs− Hoveyda-type catalyst for olefin metathesis to FhuA. This artificial metathease was then used in various chemical transformations, ranging from polymerizations (ring-opening metathesis polymerization) to enzymatic cascades involving cross-metathesis.Ultimately, we generated a catalytically active membrane by copolymerizing FhuA and pyrrole. The resulting biohybrid material was then equipped with the Grubbs−Hoveyda-type...

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Section: Fhua In Nanobiotechnology and Materials Designmentioning

confidence: 99%

Section: Fhua As a Protein Scaffold For Metal Catalystsmentioning

confidence: 99%

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FhuA: From Iron-Transporting Transmembrane Protein to Versatile Scaffolds through Protein Engineering

et al. 2023

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How to Characterize the Protein Structure and Polymer Conformation in Protein‐Polymer Conjugates – a Perspective

Mathieu-Gaedke

Böker

Glebe

2023

Macro Chemistry & Physics

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Proteins, having miscellaneous properties perfected by nature over millions of years, are attractive for many biomedical and industrial applications. The exploitation of proteins offers great opportunities to increase, among others, the selectivity, biocompatibility, and sustainability of artificial materials. However, the stability of proteins is limited and exposure to conditions like heat or organic solvents often leads to denaturation or aggregation. The modification with synthetic polymers is a powerful possibility to increase the stability of proteins. Throughout the last few years, various methods have been established to characterize obtained conjugates regarding the conjugation efficiency, protein stability, their self-assembly behavior, and beyond. Nevertheless, it is still very challenging to determine if the protein structure is unaltered in the biohybrid materials and how the polymer chains arrange around the protein surface. Here, an overview of different analysis techniques is presented, their applicability and feasibility are discussed, and current literature examples are provided. The focus of this Perspective lies on nuclear magnetic resonance spectroscopy, size exclusion chromatography coupled with multi-angle laser light scattering, analytical ultracentrifugation, and small-angle scattering techniques as these methods shed light into the structure of proteins in conjugates and the polymer conformation around proteins.

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Refolding and biophysical characterization of the Caulobacter crescentus copper resistance protein, PcoB: An outer membrane protein containing an intrinsically disordered domain

Hennaux

Kohchtali

Bâlon

et al. 2022

Biochimica et Biophysica Acta (BBA) - Biomembranes

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Protein Nanopore Membranes Prepared by a Simple Langmuir–Schaefer Approach

Cited by 3 publications

References 61 publications

FhuA: From Iron-Transporting Transmembrane Protein to Versatile Scaffolds through Protein Engineering

FhuA: From Iron-Transporting Transmembrane Protein to Versatile Scaffolds through Protein Engineering

How to Characterize the Protein Structure and Polymer Conformation in Protein‐Polymer Conjugates – a Perspective

Refolding and biophysical characterization of the Caulobacter crescentus copper resistance protein, PcoB: An outer membrane protein containing an intrinsically disordered domain

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