Mischa Zelzer

Next-generation peptide nanomaterials: molecular networks, interfaces and supramolecular functionality

With improved understanding of the design rules for self-assembling peptides, new challenges will be faced to incorporate these materials into dynamic systems of higher complexity and functionality. In this highlight article we discuss very recent advances in these areas. Three areas are covered: (i) molecular networks based on peptides and their interactions including (bio-) catalytically driven systems; (ii) supramolecular functionality, both in the context of biological and nanotechnology applications; (iii) approaches to effectively interface peptides with synthetic and biological materials. We also discuss challenges and opportunities for the design of a new generation of peptide nanomaterials for the next decade.

Enzyme responsive materials: design strategies and future developments

Enzyme responsive materials (ERMs) are a class of stimuli responsive materials with broad application potential in biological settings. This review highlights current and potential future design strategies for ERMs and provides an overview of the present state of the art in the area.

Conducting nanofibers and organogels derived from the self-assembly of tetrathiafulvalene-appended dipeptides

We demonstrate the nonaqueous self-assembly of a low-molecular-mass organic gelator based on an electroactive p-type tetrathiafulvalene (TTF)-dipeptide bioconjugate. We show that a TTF moiety appended with diphenylalanine amide derivative (TTF-FF-NH2) self-assembles into one-dimensional nanofibers that further lead to the formation of self-supporting organogels in chloroform and ethyl acetate. Upon doping of the gels with electron acceptors (TCNQ/iodine vapor), stable two-component charge transfer gels are produced in chloroform and ethyl acetate. These gels are characterized by various spectroscopy (UV-vis-NIR, FTIR, and CD), microscopy (AFM and TEM), rheology, and cyclic voltammetry techniques. Furthermore, conductivity measurements performed on TTF-FF-NH2 xerogel nanofiber networks formed between gold electrodes on a glass surface indicate that these nanofibers show a remarkable enhancement in the conductivity after doping with TCNQ.

Cooperative Self-Assembly of Peptide Gelators and Proteins

Molecular self-assembly provides a versatile route for the production of nanoscale materials for medical and technological applications. Herein, we demonstrate that the cooperative self-assembly of amphiphilic small molecules and proteins can have drastic effects on supramolecular nanostructuring of resulting materials. We report that mesoscale, fractal-like clusters of proteins form at concentrations that are orders of magnitude lower compared to those usually associated with molecular crowding at room temperature. These protein clusters have pronounced effects on the molecular self-assembly of aromatic peptide amphiphiles (fluorenylmethoxycarbonyl- dipeptides), resulting in a reversal of chiral organization and enhanced order through templating and binding. Moreover, the morphological and mechanical properties of the resultant nanostructured gels can be controlled by the cooperative self-assembly of peptides and protein fractal clusters, having implications for biomedical applications where proteins and peptides are both present. In addition, fundamental insights into cooperative interplay of molecular interactions and confinement by clusters of chiral macromolecules is relevant to gaining understanding of the molecular mechanisms of relevance to the origin of life and development of synthetic mimics of living systems.

Influence of the plasma sheath on plasma polymer deposition in advance of a mask and down pores

Plasma species that form plasma polymer deposits readily penetrate through small openings and are therefore well suited to coat the interior of porous objects. Here, we show how the size of the cross section of square channels influences the penetration of active species from a hexane plasma and how it affects the formation of surface chemical gradients in the interior of these model pores. WCA mapping and ToF-SIMS imaging are used to visualize the plasma polymer deposit in the interior of the model pores and demonstrate that a strong dependence of the wettability gradient profile only exists up to a channel cross section of about 1 mm. XPS data allow us to calculate a deposition rate of plasma polymerized hexane (ppHex) at discrete positions on the surface and show that the deposition rate of ppHex is reduced by the presence of the mask up to a distance of 16 mm in advance of the channel opening. A strong dependence of the ppHex deposition rate on the cross-section of the channels is found within the first 2 mm in front of the pore opening. An estimation of the sheath thickness suggests that this effect can be attributed to the plasma sheath that perturbs the plasma in front of the pores. Plasma mass spectrometry allows us to identify the nature of the plasma species penetrating from the plasma through the pores and shows that no negatively charged ions are able to penetrate through the small channels. Neutral and positively charged species penetrate several millimeters down the channels and both species are therefore likely to contribute to the formation of the deposit on the sample. In addition, the formation of positively charged higher molecular mass hexane fragments is observed in the gas phase, demonstrating the likelihood of neutral-positive reactions in the plasma.

Responsive cell-material interfaces.

Major design aspects for novel biomaterials are driven by the desire to mimic more varied and complex properties of a natural cellular environment with man-made materials. The development of stimulus responsive materials makes considerable contributions to the effort to incorporate dynamic and reversible elements into a biomaterial. This is particularly challenging for cell-material interactions that occur at an interface (biointerfaces); however, the design of responsive biointerfaces also presents opportunities in a variety of applications in biomedical research and regenerative medicine. This review will identify the requirements imposed on a responsive biointerface and use recent examples to demonstrate how some of these requirements have been met. Finally, the next steps in the development of more complex biomaterial interfaces, including multiple stimuli-responsive surfaces, surfaces of 3D objects and interactive biointerfaces will be discussed.

Phosphatase responsive peptide surfaces

The development of interactive surfaces able to respond to biological cues is of interest for the development of next generation biomaterials. We report the design, synthesis and characterisation of an enzyme responsive peptide based surface whose chemical properties change upon catalytic action of alkaline phosphatase (AP). AP is a membrane-anchored enzyme involved in osteogenesis (bone formation), making it a suitable biomolecule to facilitate interactivity between the cell and the biomaterial surface. Surface analysis is used to follow dephosphorylation and a phosphate assay is used to determine the amount of phosphate removed by the enzyme. This analysis reveals significant differences in the dephosphorylation rate at the surface compared to that in solution. The ability of the surface to respond to native enzymes expressed by mesenchymal stem cells (MSCs) was indirectly explored by assessing the response of the cells to phosphorylated, non-phosphorylated and enzymatically dephosphorylated surfaces. No differences were found between the surfaces, suggesting that cell expressed enzymes are able to dephosphorylate the peptide surfaces rapidly. This work presents the first phosphatase responsive surfaces whose phosphorylation state can be altered by native enzymes provided by the cells.

Dynamic Surfaces for the Study of Mesenchymal Stem Cell Growth through Adhesion Regulation

Out of their niche environment, adult stem cells, such as mesenchymal stem cells (MSCs), spontaneously differentiate. This makes both studying these important regenerative cells and growing large numbers of stem cells for clinical use challenging. Traditional cell culture techniques have fallen short of meeting this challenge, but materials science offers hope. In this study, we have used emerging rules of managing adhesion/cytoskeletal balance to prolong MSC cultures by fabricating controllable nanoscale cell interfaces using immobilized peptides that may be enzymatically activated to change their function. The surfaces can be altered (activated) at will to tip adhesion/cytoskeletal balance and initiate differentiation, hence better informing biological mechanisms of stem cell growth. Tools that are able to investigate the stem cell phenotype are important. While large phenotypical differences, such as the difference between an adipocyte and an osteoblast, are now better understood, the far more subtle differences between fibroblasts and MSCs are much harder to dissect. The development of technologies able to dynamically navigate small differences in adhesion are critical in the race to provide regenerative strategies using stem cells.

Development and validation of a fluorescence method to follow the build-up of short peptide sequences on solid 2D surfaces.

The modification of material surfaces with short peptide sequences has become an essential step in many biotechnological and biomedical applications. Due to their simple architecture compared to more complex 3D substrates, 2D surfaces are of particular interest for high throughput applications and as model surfaces for dynamic or responsive surface modifications. The decoration of these surfaces with peptides is commonly accomplished by synthesizing the peptide first and subsequently transferring it onto the surface of the substrate. Recently, several procedures have been described for the synthesis of peptides directly onto a 2D surface, thereby simplifying and accelerating the modification of flat surfaces with peptides. However, the wider use of these techniques requires a routine method to monitor the amino acid build-up on the surface. Here, we describe a fast, inexpensive and nondestructive fluorescence based method which is readily accessible to follow the amino acid build-up on solid 2D samples.

Hippocampal cell response to substrates with surface chemistry gradients.

Surface chemical gradients are valuable tools for the high-throughput screening of cell-surface interactions. However, it has yet to be shown if biological data obtained from gradient surfaces are transferable to substrates with uniform properties. To explore this question, the response of hippocampal neurons to three different sample formats was compared. We fabricated samples of uniform surface wettability and samples with a linear or radial gradient in surface wettability by depositing plasma-polymerized hexane (hydrophobic) on oxygen-etched glass (hydrophilic). Differences in cell density, growth and viability of the neural cultures are found between the uniform and the gradient samples. The nature of the gradient (linear or radial) has only a small effect on the cell density of adhered hippocampal neurons.