Engineering of the E. coli Outer Membrane Protein FhuA to overcome the Hydrophobic Mismatch in Thick Polymeric Membranes

Muhammad, Noor; Dworeck, Tamara; Fioroni, Marco; Schwaneberg, Ulrich

doi:10.1186/1477-3155-9-8

Cited by 48 publications

(40 citation statements)

References 37 publications

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“…This newly acquired metal binding ability stabilizes the helical structure of the protein, and increased the binding and lysis ability of the protein to the membrane (181). Longer transmembrane regions were also engineered for the β-barrel membrane protein FhuA to match the hydrophobic cores of thick polymeric membranes, with the goal for targeted drug delivery (182). …”

Section: Beyond Structure Prediction: Ensemble Properties Proteinmentioning

confidence: 99%

Computational studies of membrane proteins: Models and predictions for biological understanding

Liang¹,

Naveed²,

Jimenez-Morales³

et al. 2012

Biochimica et Biophysica Acta (BBA) - Biomembranes

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We discuss recent progresses in computational studies of membrane proteins based on physical models with parameters derived from bioinformatics analysis. We describe computational identification of membrane proteins and prediction of their topology from sequence, discovery of sequence and spatial motifs, and implications of these discoveries. The detection of evolutionary signal for understanding the substitution pattern of residues in the TM segments and for sequence alignment are also discussed. We further discuss empirical potential functions for energetics of inserting residues in the TM domain, for interactions between TM helices or strands, and their applications in predicting lipid-facing surfaces of the TM domain. Recent progresses in structure predictions of membrane proteins are also reviewed, with further discussions on calculation of ensemble properties such as melting temperature based on simplified state space model. Additional topics include prediction of oligomerization state of membrane proteins, identification of the interfaces for protein-protein interactions, and design of membrane proteins.

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Section: Beyond Structure Prediction: Ensemble Properties Proteinmentioning

confidence: 99%

Computational studies of membrane proteins: Models and predictions for biological understanding

Liang¹,

Naveed²,

Jimenez-Morales³

et al. 2012

Biochimica et Biophysica Acta (BBA) - Biomembranes

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“…However, their successful insertion and functionality has been reported in thick polymer membranes of poly(2‐methyl‐2‐oxazoline)‐ block ‐poly(dimethylsiloxane)‐ block ‐poly(2‐methyl‐2‐oxazoline) (PMOXA–PDMS–PMOXA), and this result was explained as being favored by the compressibility of the polymer membrane, or the polydispersity of the polymers along with segregation of the shorter chains in the vicinity of the proteins . A strategy to match the length of the hydrophobic domains of synthetic membranes with the lengths of the membrane proteins is to extend the hydrophobic region of the protein, as reported for the outer membrane protein FhuA . However, this strategy is not necessary when the synthetic membrane has sufficient flexibility, as is shown for the insertion of various channel proteins and biopores (OmpF, alamethicin, α‐haemolysin, LamB, Tsx, FhuA, and AqpZ) in PMOXA–PDMS–PMOXA polymersomes .…”

Section: Introductionmentioning

confidence: 99%

Does Membrane Thickness Affect the Transport of Selective Ions Mediated by Ionophores in Synthetic Membranes?

Lomora

Dinu

Itel

et al. 2015

Macromol. Rapid Commun.

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Biomimetic polymer nanocompartments (polymersomes) with preserved architecture and ion-selective membrane permeability represent cutting-edge mimics of cellular compartmentalization. Here it is studied whether the membrane thickness affects the functionality of ionophores in respect to the transport of Ca ions in synthetic membranes of polymersomes, which are up to 2.6 times thicker than lipid membranes (5 nm). Selective permeability toward calcium ions is achieved by proper insertion of ionomycin, and demonstrated by using specific fluorescence markers encapsulated in their inner cavities. Preservation of polymersome architecture is shown by a combination of light scattering, transmission electron microscopy, and fluorescence spectroscopy. By using a combination of stopped-flow and fluorescence spectroscopy, it is shown that ionomycin can function and transport calcium ions across polymer membranes with thicknesses in the range 10.7-13.4 nm (7.1-8.9 times larger than the size of the ionophore). Thicker membranes induce a decrease in transport, but do not block it due to the intrinsic flexibility of these synthetic membranes. The design of ion selective biomimetic nanocompartments represents a new path toward the development of cellular ion nanosensors and nano-reactors, in which calcium sensitive biomacromolecules can be triggered for specific biological functions.

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“…To our knowledge this is the first time, besides our previous study [16], a channel protein was specifically engineered to modify its geometry.…”

Section: Discussionmentioning

confidence: 95%

Engineering of an E. coli outer membrane protein FhuA with increased channel diameter

2011

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BackgroundChannel proteins like FhuA can be an alternative to artificial chemically synthesized nanopores. To reach such goals, channel proteins must be flexible enough to be modified in their geometry, i.e. length and diameter. As continuation of a previous study in which we addressed the lengthening of the channel, here we report the increasing of the channel diameter by genetic engineering.ResultsThe FhuA Δ1-159 diameter increase has been obtained by doubling the amino acid sequence of the first two N-terminal β-strands, resulting in variant FhuA Δ1-159 Exp. The total number of β-strands increased from 22 to 24 and the channel surface area is expected to increase by ~16%. The secondary structure analysis by circular dichroism (CD) spectroscopy shows a high β-sheet content, suggesting the correct folding of FhuA Δ1-159 Exp. To further prove the FhuA Δ1-159 Exp channel functionality, kinetic measurement using the HRP-TMB assay (HRP = Horse Radish Peroxidase, TMB = 3,3',5,5'-tetramethylbenzidine) were conducted. The results indicated a 17% faster diffusion kinetic for FhuA Δ1-159 Exp as compared to FhuA Δ1-159, well correlated to the expected channel surface area increase of ~16%.ConclusionIn this study using a simple "semi rational" approach the FhuA Δ1-159 diameter was enlarged. By combining the actual results with the previous ones on the FhuA Δ1-159 lengthening a new set of synthetic nanochannels with desired lengths and diameters can be produced, broadening the FhuA Δ1-159 applications. As large scale protein production is possible our approach can give a contribution to nanochannel industrial applications.

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Engineering of the E. coli Outer Membrane Protein FhuA to overcome the Hydrophobic Mismatch in Thick Polymeric Membranes

Cited by 48 publications

References 37 publications

Computational studies of membrane proteins: Models and predictions for biological understanding

Computational studies of membrane proteins: Models and predictions for biological understanding

Does Membrane Thickness Affect the Transport of Selective Ions Mediated by Ionophores in Synthetic Membranes?

Engineering of an E. coli outer membrane protein FhuA with increased channel diameter

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