A Designed Biosurfactant Protein for Switchable Foam Control

Middelberg, Anton P. J.; Dimitrijev-Dwyer, Mirjana

doi:10.1002/cphc.201100082

Cited by 53 publications

(88 citation statements)

References 34 publications

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“…It is expected that DAMP1 will behave similarly. Like AM1 [19], DAMP1 is largely unstructured in bulk (see the electronic supplementary material, figure S1), whereas DAMP4 forms a very stable four-helix bundle [15].…”

Section: Introductionmentioning

confidence: 99%

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Insights into the role of protein molecule size and structure on interfacial properties using designed sequences

Dwyer

James

et al. 2013

J. R. Soc. Interface.

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Mixtures of a large, structured protein with a smaller, unstructured component are inherently complex and hard to characterize at interfaces, leading to difficulties in understanding their interfacial behaviours and, therefore, formulation optimization. Here, we investigated interfacial properties of such a mixed system. Simplicity was achieved using designed sequences in which chemical differences had been eliminated to isolate the effect of molecular size and structure, namely a short unstructured peptide (DAMP1) and its longer structured protein concatamer (DAMP4). Interfacial tension measurements suggested that the size and bulk structuring of the larger molecule led to much slower adsorption kinetics. Neutron reflectometry at equilibrium revealed that both molecules adsorbed as a monolayer to the air -water interface (indicating unfolding of DAMP4 to give a chain of four connected DAMP1 molecules), with a concentration ratio equal to that in the bulk. This suggests the overall free energy of adsorption is equal despite differences in size and bulk structure. At small interfacial extensional strains, only molecule packing influenced the stress response. At larger strains, the effect of size became apparent, with DAMP4 registering a higher stress response and interfacial elasticity. When both components were present at the interface, most stress-dissipating movement was achieved by DAMP1. This work thus provides insights into the role of proteins' molecular size and structure on their interfacial properties, and the designed sequences introduced here can serve as effective tools for interfacial studies of proteins and polymers.

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Section: Introductionmentioning

confidence: 99%

“…15 Å ) [18]. AM1, and subsequently DAMP4 and DAMP1, are designed with hydrophobic and hydrophilic residues placed strategically to drive helical interfacial structuring [15,17]. DAMP4 and DAMP1 are, therefore, expected to form a similar monolayer structure.…”

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confidence: 99%

Insights into the role of protein molecule size and structure on interfacial properties using designed sequences

Dwyer

James

et al. 2013

J. R. Soc. Interface.

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“…Targeted design approaches such as those outlined in Section 1.2 may be utilised to give peptide and concatemer sequences that will fold into stable structures for expression. [316][317][318] Both theoretical considerations and a lack of examples in the literature suggest tandem expression of 39 highly cationic or anionic peptides to be unfavourable. Expression of these highly charged sequences is constrained by charge destabilisation preventing stable folding within the cell, and potential host toxicity in the case of cationic sequences.…”

Section: Biological Expressionmentioning

confidence: 99%

“…The majority of published methods for the purification of expressed peptides utilise reversed phase, affinity or ion-exchange chromatography, either in single or sequential steps. 274,306,311,312,317,[341][342][343] This is due to the high level of purity attained with these methods as well as technique simplicity and adaptability for varied peptides. While these methods are effective at the laboratory scale for early peptide characterisation, chromatography-based approaches add significant cost to overall peptide production and are impractical for large-scale processes.…”

Section: Biological Expressionmentioning

confidence: 99%

“…DAMP4 was then designed as a tetrameric coiled-coil concatemer of the AM1 sequence separated by acid-cleavable linkers which was recovered using a chromatographic approach. 317 More recent work by the same group has produced a chromatography-free process for recovery of "industrial grade" DAMP4. 346 However, deamidation of asparagine residues occurred during cleavage of the expressed protein that affected the interfacial properties of the recovered peptide.…”

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Bioproduction and self-assembly of designer peptides

Fletcher¹

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Self-assembly is a powerful method for producing controlled morphologies at the nanometre scale. Peptides are biomolecules capable of self-assembly and have the potential for use in a variety of applications such as emulsion and foam stabilisation, wound healing and drug delivery. The investigation of peptide sequence-structure relationships is a rapidly advancing field of study, which in many cases now allows the targeted design of self-assembling peptides.While self-assembling peptides show potential in several fields, their large-scale adoption is limited by the high production expenses associated with conventional solid-phase synthesis. A potentially cheaper and more renewable approach to peptide production is bacterial expression. However, the bioproduction of peptides is a non-trivial process, and generally involves the expression of peptides as part of a fusion protein in which the target peptide is only a small portion of the expressed product. An alternate approach involves peptide concatemers, in which the target peptide makes up the majority of the expressed construct.The work reported in this thesis focused on the design, characterisation and bioproduction of self-assembling α-helical peptides. It was particularly focussed on the development of amphiphillic α-helical peptides with applications as surfactants and hydrogelators. The aim of this work was to further the field of de novo peptide design, while also investigating an approach for peptide bioproduction. This work aimed to combine the requirements for peptide bioproduction with end-use functionality.Chapter 2 details successful bioproduction of an anionic helical surfactant peptide EDP-11, as part of a charge-paired heteroconcatemer with the cationic expression partner RDP-4. The method utilised designed assembly of the constituent α-helical peptides to generate a stably folded coiled-coil concatemer. The polypeptide sequence was further optimized for molecular charge, hydropathy and predicted protease resistance. This process allowed expression of a soluble concatemer that accumulated to high levels (22% of total protein) in E. coli. The expressed concatemer possessed extreme stability to heat and proteases, allowing isolation by simple heat and pH precipitation, yielding concatemer at 130 mg per gram of dry cell weight and >99% purity. Key parameters used in designing the heteroconcatemer were then compared to those of all open reading frames of several reference ii proteomes in an attempt to gain insight into the mechanistic basis for the high stability of the designed miniprotein.Following bioproduction using this concatemer approach, further processing was required to produce monomeric peptides for use in surfactant applications. The design of acid-cleavable aspartate-proline sites within the concatemer sequence allowed for simple heat-and acid-mediated cleavage to give constituent peptides.Chapter 3 details characterization of cleavage of the expressed concatemer by coupled liquid chromatography-mass spectrometry and includes mod...

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Design of Stimuli‐Responsive Peptides and Proteins

Yang

Gerstweiler

et al. 2022

Adv Funct Materials

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Stimuli‐responsive peptides and proteins are an exciting class of smart biomaterials for various applications and have received significant attention over the past decades. A wide variety of stimuli such as temperature, pH, ions, enzymes, magnetic field, redox, etc., are explored. This article provides a review of five intensively studied types of stimuli‐responsive peptides and proteins, their design principles and applications, including temperature‐, pH‐, light‐, metal ion‐, and enzyme‐responsive with an emphasis on the key design concepts and switch function. Moreover, typical examples of their applications are discussed to provide a better understanding of the design concept and underlying methodology. This review will facilitate and inspire future innovation toward new peptide‐ and protein‐based materials and their diverse applications.

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A Designed Biosurfactant Protein for Switchable Foam Control

Cited by 53 publications

References 34 publications

Insights into the role of protein molecule size and structure on interfacial properties using designed sequences

Insights into the role of protein molecule size and structure on interfacial properties using designed sequences

Bioproduction and self-assembly of designer peptides

Design of Stimuli‐Responsive Peptides and Proteins

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