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Self-assembled peptides: from nanostructure to bioactivity

Scientific meeting

Starts:

October
242016

09:00

Ends:

October
252016

17:00

Location

Kavli Royal Society Centre, Chicheley Hall, Newport Pagnell, Buckinghamshire, MK16 9JJ

Overview

Theo Murphy international scientific discussion meeting organised by Professor Ian Hamley, Professor Dek Woolfson, Professor Louise Serpell, Dr Alberto Saiani and Professor Raffaele Mezzenga.

Self assembly of LACQCL, a fragment of betalactoglobulin forming amyloids involved in capturing heavy metal ions from water. Copyright Professor Raffaele Mezzenga

This meeting discussed exciting emerging themes in understanding and controlling the self-assembly of peptides to create unique nanostructures, and to enable targeted and enhanced bioactivity from the presentation of the peptide motifs. It covered several classes of self-assembling peptides and related molecules including amyloid peptides, lipopeptides and coiled coil peptides.

"This meeting presents an unparalleled opportunity for conferees to discuss peptide self-assembly with world leaders in the field in an open and intimate format and the superb setting of the Royal Society's Chicheley Hall" Professor Dek Woolfson

Attending the event

This meeting has taken place.

Recorded audio of the presentations are available for each talk below. Meeting papers will be published in a future issue of Interface Focus.

For further information please contact the scientific programmes team.

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Schedule of talks

24 October

09:00-12:40

Session 1

5 talks Show detail Hide detail

Chairs

Professor Ian Hamley, University of Reading, UK

09:05-09:45 Designer self-assembling peptides:nanofibers, nanotubes and dynamic behaviours

Professor Shuguang Zhang, Center for Bits and Atoms, MIT, USA

Abstract

Short peptides that are made of natural amino acids were never seriously considered as useful materials 25 years ago. However, the discovery of a class of self-assembling peptides in yeast that spontaneously undergo self-organization into well-ordered structures resulted in a conceptual change. Since then diverse classes of short peptides have been invented with broad applications including 3D tissue cell culture, reparative and regenerative medicine, tissue engineering, slow and sustained drug release, stabilization of membrane proteins for develop nanobiotechnology and molecular devices. Molecular design using short peptides as new materials will likely play increasingly important role in nanoscience, nanotechnology, nanobiotechnology and nanomedicine.

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09:45-10:25 The diversity and utility of amyloid fibrils

Professor Louise Serpell, University of Sussex, UK

Abstract

Amyloidogenic peptides are well known for their involvement in diseases such as Diabetes type 2 and Alzheimer’s disease. However, more recently, amyloid fibrils have been shown to provide scaffolding and protection as functional materials in a range of organisms from bacteria to humans. These roles highlight the incredible tensile strength of the cross-beta amyloid architecture. Many sequences are able to self-assemble to form amyloid with a cross-beta core and the group have explored the contribution of the amino-acid side chains to amyloidogenicity and the amyloid architecture using X-ray fibre diffraction and electron microscopy. The group have described a 12mer peptide that forms fibrous crystals, a seven residue peptide originating from alpha-synuclein that has the ability to form nanotubes and a range of penta- and hexapeptides forming different morphologies. This work provides a wide range of peptides that may be exploited as fibrous bionanomaterials. These fibrils provide a scaffold upon which functional groups may be added, or templated assembly may be performed. This talk will present recent work on developing fluorescent nanotubes, silica-nanowires and peptide catalysts.

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10:25-10:40 Coffee/Tea

10:40-11:20 Localising structure with peptides and peptoids

Dr K. H. Aaron Lau, University of Strathclyde, Glasgow, UK

Abstract

The group present recent results using peptidomimetic molecules to define the spatial arrangements of chemical groups and nanostructures on material surfaces, as steps towards the mimicry of natural biointerfaces and their function. In a first example, the juxtaposition on the molecular scale of chemical groups is defined using the primary sequence of peptidomimetic poly(N-substituted glycine) “peptoids”. By modifying the spatial separation between oppositely charged chemical groups and their sequence order, the electrostatic interaction between proteins and a peptoid-coated surface can be controlled. In a second example, proteases are coated on a surface to enable the reverse hydrolysis of peptide molecular gelator precursors. By tuning the conditions of the surface preparation, self-assembled peptide nanofibers may be observed in the bulk solution or preferentially localized on the surface. The peptidomimetic systems presented are convenient for incorporating biochemical functionality and for organizing these groups on both the molecular and nanostructural length-scales. The localization of enzymatic activity especially augurs well for generating dynamic and adaptive biointerfaces.

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11:20-12:00 Engineering Peptide Display for Self-assembly and Interactions with Cells

Dr Helena Azevedo, Queen Mary University of London, UK

Abstract

The possibility of displaying biomolecular functionalities (e.g. small peptide molecules) with spatiotemporal control for triggering macromolecular (e.g. polysacharides, enzymes or cell surface receptors) recognition events is an important goal in biomaterials engineering. Progress in this area will enable the design of defined substrates for cell culture studies in 2- or 3-D or the development of smart drug delivery carriers. This talk will present our recent efforts on peptide engineering to trigger their self-assembly into self-supporting gels and micellar nanocarriers or onto surfaces.

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12:00-12:40 Drug Delivery via Cell Membrane Fusion using Lipopeptide Modified Liposomes

Professor Alexander Kros, Leiden Institute of Chemistry, Netherlands

Abstract

Efficient delivery of drugs to living cells is still a major challenge. Currently, most methods rely on the endocytotic pathway resulting in low delivery efficiency due to limited endosomal escape and/or degradation in lysosomes. Here, we report a new method for direct drug delivery into the cytosol of live cells in vitro and vivo utilizing targeted membrane fusion between liposomes and live cells. A pair of complementary coiled coil lipopeptides was embedded in the lipid bilayer of liposomes and cell membranes respectively, resulting in targeted membrane fusion with concomitant release of liposome encapsulated cargo including fluorescent dyes and the cytotoxic drug doxorubicin. Using a wide spectrum of endocytosis inhibitors and endosome trackers we demonstrate that the major site of cargo release is at the plasma membrane. This method thus allows for the quick and efficient delivery of drugs and is expected to have many in-vitro, ex-vivo and in-vivo applications.

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12:40-13:40 Lunch

13:40-17:30

Session 2

5 talks Show detail Hide detail

Chairs

Professor Louise Serpell, University of Sussex, UK

13:40-14:20 Self-assembly and immunity

Professor Gerard Wong, Bioengineering Department, Chemistry & Biochemistry Department, CNSI, UCLA, USA

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14:20-15:00 Peptide Conjugates: From Self-Assembly to Bioactivity

Professor Ian Hamley, University of Reading, UK

Abstract

Self-assembling peptides and their conjugates offer exceptional potential in nanomedicine. This talk will present some of the group's recent work on nanoscale assembled peptides and their conjugates, focussing on lipopeptides. Examples from recent work on self-assembling lipopeptides (peptides attached N- terminally to palmitoyl, hexadecyl, lipid chains) will be outlined. The group's focus is to investigate potential relationships between self-assembly and bioactivity, in particular in the fields of regenerative medicine, antimicrobial systems, and immune therapies.

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15:00-15:30 Coffee/Tea

15:30-16:10 The Effect of Glycation on the Triple Helix Forming Properties of Collagen Model Peptides

Professor Margaret Brimble, University of Auckland, New Zealand

Abstract

Advanced glycation endproducts (AGEs) are a family of unnatural amino acids that result from non-enzymatic reactions between reducing sugars and amino acid side-chains in proteins. These post-translational modifications mainly involve reactions at lysine and arginine residues and give rise to a large number of structurally diverse compounds ranging from simple alkylation to more complex heterocycles and cross-linked structures. The formation and accumulation of AGEs in the body is thought to play a significant role in the pathogenesis of many debilitating diseases including diabetes and age-related neurodegenerative diseases, such as Alzheimer’s disease.

The precise molecular mechanisms by which AGEs influence the progression of these diseases are not clear, and there remains a need to develop new chemical tools to investigate them. Collagen is the most abundant protein in vertebrates and its long-lived nature makes it susceptible to AGE formation.  Collagen is therefore the most relevant protein to study when investigating AGEs.

The synthesis of several AGE-modified amino acid building blocks and their site-specific incorporation into collagen model peptide (CMP) sequences using SPPS is described. The AGE-CMPs were shown to form triple helices in solution demonstrating their potential as models for investigating AGE-modified collagens. Cross-linking of proteins by AGEs causes a host of pathological conditions but their exact roles are unknown. Cross-linking lysyl AGEs were also synthesized and incorporated into two types of collagen peptides. The utility of these cross-linked peptides for biochemical investigations was demonstrated by proteolysis studies and circular dichroism.

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16:10-16:50 Rational Hydrogel Design Enabled by Structural Insight into Hydrogelation of Self-Assembled Materials

Professor Bradley Nilsson, University of Rochester, USA

Abstract

The self-assembly of peptides and proteins into cross-amyloid structures is a defining characteristic of amyloid pathologies, including Alzheimer’s disease, Parkinson’s disease, type 2 diabetes and prion encephalopathies. Amyloid protein assemblies also exist as evolutionarily conserved motifs with defined biological function. There is growing interest in exploiting self-assembled peptides in the development of novel functional structures with applications in biomedicine, energy, and materials. Self-assembled peptides that form emergent hydrogel networks have been of special significance for applications in tissue engineering and regenerative medicine. Herein, efforts to bridge the gap between empiricism and rational design in the development of self-assembled hydrogels derived from functionalized amino acids will be discussed. Studies focused on understanding the self-assembly mechanisms and the packing structure of functionalized phenylalanine assemblies have provided insight that enables the use of these agents to create functional supramolecular hydrogels. These low molecular weight hydrogels possess properties similar to those exhibited by materials derived from more expensive peptide-based systems.

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16:50-17:30 Directed discovery of peptide nanostructures

Professor Rein V. Ulijn, CUNY Advanced Science Research Center, USA

Abstract

It is the group’s aim to develop functional nanostructures that mimic the ability of living systems to sense, adapt, convert energy and respond to new situations but are drastically simplified, robust and functional. The design and selection of suitable self-assembling sequences is, however, challenging due to the vast combinatorial space available. This talk will report on directed discovery methodology that allows the peptide sequence space to be searched for self-assembling structures using computation and experiment. An additional challenge in mimicry of living materials is that they actively (rather than passively) respond to new situations. This talk will report on the progress of developing such non-equilibrium nanostructures. A number of examples of functional bio-inspired nanostructures will be discussed with applications in design of responsive biomaterials and tunable emulsifiers.

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25 October

09:00-12:45

Session 3

5 talks Show detail Hide detail

Chairs

Professor Dek Woolfson, University of Bristol, UK

09:00-09:40 Autoimmunity and neurodegeneration: the mimicry pathway from aberrant N-glucosylated peptides to specific protein antigens

Professor Anna Maria Papini, PeptLab, University of Florence and University of Cergy Pontoise

Abstract

In autoimmune diseases, it has been proposed that exogenous "molecular triggers", i.e., specific non-self-antigens accompanying infectious agents, may disrupt the control of adaptive immune system resulting in serious pathologies. The aetiology of multiple sclerosis (MS) remains unclear. However, epidemiologic data suggest that exposure to infectious agents may be associated with increased MS risk and that progression may be linked to exogenous, bacterially-derived, antigenic molecules mimicking mammalian cell surface glycoconjugates triggering autoimmune responses. Previously, antibodies specific to a gluco-asparagine (N-Glc) glycopeptide, CSF114(N-Glc), were identified in sera of an MS patient subpopulation. Since the human glycoproteome repertoire lacks this uniquely modified amino acid, the group turned its attention to bacteria, i.e., Haemophilus influenzae, expressing cell-surface adhesins including N-Glc, to establish connection between H. influenzae infection and MS. The group exploited the biosynthetic machinery from H. influenzae opportunistic pathogens (and the homologous enzymes from A. pleuropneumoniae) to produce a unique set of defined glucosylated adhesin proteins. Interestingly it revealed that a hyperglucosylated protein domain, based on the cell-surface adhesin HMW1A, is preferentially recognized by antibodies from sera of an MS patient subpopulation. In conclusion the hyperglucosylated adhesin is the first example of an N-glucosylated native antigen that can be considered a relevant candidate for triggering pathogenic antibodies in MS.

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09:40-10:20 Design of Structurally Defined 1D and 2D assemblies.

Professor Vincent Conticello, Emory University, Atlanta, GA USA

Abstract

Structurally defined materials on the nanometer length-scale have been historically the most challenging to rationally construct and the most difficult to structurally analyze. Sequence-specific biomolecules, i.e., proteins and nucleic acids, have advantages as design elements for construction of these types of nano-scale materials in that correlations can be drawn between sequence and higher order structure, potentially affording ordered assemblies in which functional properties can be controlled through the progression of structural hierarchy encoded at the molecular level. However, the predictable design of self-assembled structures requires precise structural control of the interfaces between peptide subunits (protomers). In contrast to the robustness of protein tertiary structure, quaternary structure has been postulated to be labile with respect to mutagenesis of residues located at the protein-protein interface. We have employed simple self-assembling peptide systems to interrogate the concept of designability of interfaces within the structural context of nanotubes and nanosheets. These peptide systems provide a framework for understanding how minor sequence changes in evolution can translate into very large changes in supramolecular structure, which provides significant evidence that the designability of protein interfaces is a critical consideration for control of supramolecular structure in self-assembling systems.

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10:20-10:45 Coffee/Tea

10:45-11:25 Dehydropeptide hydrogelators as new potential drug nanocarriers

Professor Paula Margarida Ferreira, University of Minho, Portugal

Abstract

Hydrogels made of small peptides, especially di- and tripeptides, are particularly attractive owing to simple synthetic procedures, chemical variability, and potential for introduction biological functionality. The gelation of this type of peptides is usually driven by the cooperative effect of several weak intermolecular interactions such as hydrogen bonding, hydrophobic and aromatic-interactions. The main limitation of peptide hydrogels for biomedical applications is their susceptibility to enzymatic hydrolysis. A well established strategy to provide peptides and proteins with proteolytic stability is the replacement of natural amino acids by non-proteinogenic analogues such as D-amino acids, ß-amino acids or dehydroamino acids.

Small peptides with dehydroamino acids residues and N-capped with bulky aromatic moieties constitute an important class of hydrogelators that resists proteolysis. The dehydropeptides building blocks can be easily obtained from the corresponding peptides with β-hydroxyamino acid residues and give hydrogels at low critical gelation concentrations. Molecular dynamic simulations showed that the conformational constraints imposed by the dehydroamino acid residues are beneficial to the peptide self-assembly process into supramolecular structures. Preliminary studies using photophysical methods showed that these materials can be used as efficient nanocarriers of several antitumoral drugs.

Hydrogels obtained from small dehydropeptides can be considered as novel nano-pharmaceuticals since they ally proteolytic stability to potential intrinsic biological activity and drug delivery capability.

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11:25-12:05 Heterotypic supramolecular assemblies of short peptidic derivatives

Professor Bing Xu, Brandeis University, Massachusetts, USA

Abstract

Supramolecular hydrogels, formed via intermolecular interactions in water, are emerging as a new type of versatile soft materials to be applied in many areas, such as biomedical applications, catalysis, food additives, and cosmetics. While most of the supramolecular hydrogels are homotypic (i.e., one type of building blocks), heterotypic supramolecular hydrogels are less explored, but may offer unique advantages. This talk discuss supramolecular hydrogels that consist of more than one type building blocks (i.e., heterotypic) to illustrate the promises and challenges of heterotypic supramolecular hydrogels as soft biomaterials. First, we discuss the driving force for producing heterotypic supramolecular hydrogels. Second, we introduce the general methods for triggering heterotypic supramolecular hydrogels. Third, we report an example of two complementary pentapeptides from a beta-sheet motif of a protein self-assemble to form beta-sheet like structures upon being mixed in water.  Although beta-sheet is a common secondary structure formed by certain segments of peptides in proteins, the isolated segments by themselves usually are unable to maintain the original secondary structures due to the lack of the conformation restriction provided by the proteins. By promoting the pentapeptides transform from alpha-helix to beta-sheet conformation, the self-assembly results in supramolecular hydrogels. This talk will illustrate a bioinspired way to generate supramolecular peptide nanofibers, a class of bioactive entities, with predefined secondary structures by a rational design that uses protein structures as the blueprint.

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12:05-12:45 SNARE mimicking peptides for membrane fusion

Professor Ulf Diederichsen, Georg-August-Universitat, Germany

Abstract

Membrane fusion in case of synaptic transmission is triggered by fusion proteins like SNARE (Soluble N-ethylmaleimide-Sensitive Factor Attachment Protein Receptor) proteins. A coiled-coil four-helix bundle is formed between SNARE proteins syntaxin-1A and SNAP-25 residing in the plasma membrane and the SNARE protein synaptobrevin residing in the membrane of synaptic vesicles, forcing the two merging membranes in close proximity. The precise mechanism of SNARE mediated membrane fusion, e.g. the role of transmembrane domains of synaptobrevin (Syb) and syntaxin-1A (Sx), is still under debate. Therefore, fusion experiments are described using vesicles reconstituted with artificial SNARE mimicking model systems, thereby, simplifying the SNARE assembly reaction and allowing systematic structural variations. SNARE analogous model systems based on transmembrane peptides and covalently linked recognition motifs like coiled-coil forming peptides or peptide nucleic acids (PNAs) are described. The PNA recognition motif is especially suited to control the directionality of the recognition process using caging groups thereby controlling vesicle docking and fusion. In addition, an interdependence between the recognition process and the peptide helix transmembrane domain was postulated with respect to vesicle docking and fusion efficiency. The transmembrane peptide and especially the charge of C-terminus have significant influence on the fusion mechanism.

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12:45-13:45 Lunch

13:45-17:00

Session 4

4 talks Show detail Hide detail

Chairs

Dr Helena Azevedo, Queen Mary University of London, UK

13:45-14:10 Therapeutic peptides amyloid self-assembly: the concerns and the likelihoods to the pharmaceutical industry

Dr Ana Dos Santos, Medimmune Ltd, Cambridge, UK

Abstract

The use of peptides as therapeutics is undergoing a renewed enthusiasm owing to a notable expansion of the number of marketing approvals in the recent years. Current successes in the development of peptides as therapeutics are likely to spur additional growth in this sector. However, clinical uses of peptide-based medicines are still challenged by short in vivo life time and low stability. Chemical modifications, such as substitutions, acylation and PEGylation, have compensated some but not all of their promises. From the development point of view, the reduced size of peptides contrasted with larger biologics upsurge to different formulation hurdles, mostly towards to chemical, conformational stability and aggregation. These unique features result in challenges of attaining satisfactory chemical and physical stability through the manufacturing process and during shelf-life. Hence, this complex feature also makes them some of the most thought-provoking molecules to design, formulate and deliver. Strategies to limit the aggregation of peptides during storage are likely to benefit from the recent surge of interest in peptide fibrillation. This talk focuses on a brief overview of formulation issues and the approaches to mitigate some development obstacles associated with peptide therapeutics self-assembly and fibrillation. The presenter also emphasizes opportunities for the thoughtful application of peptide self-assembly to develop long acting peptide drugs.

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14:25-14:50 Peptide gels for slow release drug delivery and antimicrobial applications

Professor Arindam Banerjee, Indian Association for the Cultivation of Science, India

Abstract

Molecular self-association plays a pivotal role in chemical, biological and material sciences. Peptides with suitable functionalities are good candidates for assembly by using various non-covalent interactions including hydrogen bonding, pi-pi stacking, electrostatic, solvophobic and others. Under a suitable situation, a peptide can be self-assembled to form a micro/nano-fibrillar network structure occupied by a large amount of solvent molecules (water/organic solvent) and this forms a soft material called gel. It is interesting to control the assembly of designer oligopeptides to make useful gels and also to explore fascinating applications of these gels.

These gels were applied to perform a variety of functions including carriers of drugs and other biologically active molecules. Recently, a peptide-based thxiotropic, proteolytically stable hydrogel has been designed and constructed and it has shown a remarkable antibacterial properties against Gram-negative bacteria. Moreover, it has shown biocompatibility with human red blood cells and human fibroblast cells. Another interesting study demonstrates peptide based soft biomaterials for anti-cancer drug release and modulation of stiffness, drug release capacity and proteolytic stability of these hydrogels by incorporating D-amino acid residue(s).

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15:05-15:30 Coffee/Tea

15:30-16:10 Peptide inhibitory nanoparticles as a multivalent inhibitor of Aβ aggregation and potential treatment for Alzheimer’s disease

Professor David Allsop, Lancaster University, UK

Abstract

Aggregation of Amyloid-β peptide (Aβ) into small oligomers, and into the amyloid fibrils associated with senile plaques, is a key event in the pathogenesis of Alzheimer’s disease.  The group have investigated the effects of nanoliposomes decorated with the retro-inverso peptide RI-OR2-TAT (Ac-rGffvlkGrrrrqrrkkrGy-NH2) on the aggregation and toxicity of Aβ. This retro-inverted peptide has been shown to bind to Aβ monomers and to prevent their assembly into oligomers and β-pleated sheet fibrils. Remarkably low concentrations of Peptide Inhibitory NanoParticles (PINPs) were required to inhibit the formation of Aβ oligomers and fibrils in vitro, with 50% inhibition occurring at a molar ratio of ~1:2000 of liposome-bound RI-OR2-TAT to Aβ. PINPs also bound to Aβ with high affinity (Kd = 13.2 - 50 nM), rescued SHSY-5Y cells from the toxic effects of pre-aggregated Aβ, crossed an in vitro blood-brain-barrier model (hCMEC/D3 cell monolayer), entered the brains of C57/BL6 mice, and protected against memory loss in APPSWE transgenic mice in a novel object recognition test. As the most potent Aβ aggregation inhibitor that has been tested so far, the group propose to develop PINPs as a potential disease-modifying treatment for slowing progression of Alzheimer’s disease.

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16:10-16:50 Kinetics of protein aggregation

Professor Tuomas Knowles, University of Cambridge, UK

Abstract

This talk outlines the efforts by the group to explore the use of peptides and proteins as building blocks for the synthesis of microcapsules. The group use the self-assembly of polypeptide chains into nanofibrils to define the structure of these materials on the nanoscale, and exploit microfluidics to determine their micron scale morphology. Such capsules can be used for the stabilisation, storage and release of sensitive materials, in particular aggregation prone antibodies. Moreover, we explore the use of peptide self-assembly within microdroplets as the basis of active materials and demonstrate chemo-mechanical actuation in such systems.

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Self-assembled peptides: from nanostructure to bioactivity Kavli Royal Society Centre, Chicheley Hall Newport Pagnell Buckinghamshire MK16 9JJ