Obtaining an understanding, at the atomic level, of the interaction of nanomaterials with biological systems has recently become an issue of great research interest. Here we report on the molecular ...dynamics study of the translocation of fullerene C60 and its derivative C60(OH)20 across a model cell membrane (dipalmitoylphosphatidylcholine or DPPC bilayer). The simulation results indicate that, although a pristine C60 molecule can readily “jump” into the bilayer and translocate the membrane within a few milliseconds, the C60(OH)20 molecule can barely penetrate the bilayer. Indeed, the mean translocation time via diffusion for the C60(OH)20 molecule is several orders of magnitude longer than for the former. It was also determined that the two different forms of fullerenes, when adsorbed into/onto the bilayer, affected the membrane structure differently. This study offers a mechanistic explanation of that difference and for the reduced acute toxicity of functionalized fullerenes.
The fabrication of protected peptide-based hydrogels on electrode surfaces can be achieved by employing the electrochemical oxidation of hydroquinone to benzoquinone, liberating protons at the ...electrode-solution interface. The localised reduction in pH below the dipeptide gelator molecules p
initiates the neutralisation, self-assembly and formation of self-supporting hydrogels exclusively at the electrode surface. Previous examples have been on a nanometre to millimetre scale, using deposition times ranging from seconds to minutes. However, the maximum size to which these materials can grow and their subsequent mechanical properties have not yet been investigated. Here, we report the fabrication of the largest reported di- and tri-peptide based hydrogels using this electrochemical method, employing deposition times of two to five hours. To overcome the oxidation of hydroquinone in air, the fabrication process was performed under an inert nitrogen atmosphere. We show that this approach can be used to form multilayer gels, with the mechanical properties of each layer determined by gelator composition. We also describe examples where gel-to-crystal transitions and syneresis occur within the material.
•Effect of PEG spacer length was evaluated for enhanced analytical performance.•PEG-6 spacer provided enhanced electrode surface anti-fouling properties.•Optimisation of an electrochemical biosensor ...platform for monitoring protease activity.•Redox-tagged peptides with varying pegylated spacer lengths were synthesised.
Peptide-based electrochemical biosensors typically consist of a short peptide sequence, labelled with a redox reporter and modified with a thiol-containing moiety to allow immobilisation onto a gold electrode surface. A spacer is often introduced between the thiol group and the peptide with the aim of promoting enzyme accessibility as well as conferring flexibility onto the probe. Herein we report a systematic study of the effect of polyethylene glycol (PEG)-based spacer length on the performance of such biosensors in order to gain a deeper understanding of their role and optimise a peptide-based electrochemical sensor. Thus, a specific peptide endowed with varying PEG spacers (PEG-4, PEG-6, PEG-8 and PEG-12) were synthesised and interrogated by the addition of both a target enzyme (trypsin) and BSA in order to evaluate their analytical performance. An alkyl-based spacer was also assessed in order to compare the effect of the nature of the spacer. All of the proposed probes supported efficient protease detection; however, PEG-6 provided enhanced anti-fouling properties, which highlights the vital role of the spacer in the design of peptide-based probes.
A ruthenium-based mitochondrial-targeting photosensitiser that undergoes efficient cell uptake, enables the rapid catalytic conversion of Pt
prodrugs into their active Pt
counterparts, and drives the ...generation of singlet oxygen was designed. This dual mode of action drives two orthogonal cancer-cell killing mechanisms with temporal and spatial control. The designed photosensitiser was shown to elicit cell death of a panel of cancer cell lines including those showing oxaliplatin-resistance.
Thoracian barnacles rely heavily upon their ability to adhere to surfaces and are environmentally and economically important as biofouling pests. Their adhesives have unique attributes that define ...them as targets for bio-inspired adhesive development. With the aid of multi-photon and broadband coherent anti-Stokes Raman scattering microscopies, we report that the larval adhesive of barnacle cyprids is a bi-phasic system containing lipids and phosphoproteins, working synergistically to maximize adhesion to diverse surfaces under hostile conditions. Lipids, secreted first, possibly displace water from the surface interface creating a conducive environment for introduction of phosphoproteins while simultaneously modulating the spreading of the protein phase and protecting the nascent adhesive plaque from bacterial biodegradation. The two distinct phases are contained within two different granules in the cyprid cement glands, implying far greater complexity than previously recognized. Knowledge of the lipidic contribution will hopefully inspire development of novel synthetic bioadhesives and environmentally benign antifouling coatings.
Human neutrophil elastase (HNE) is a serine protease, produced by polymorphonuclear neutrophils (PMNs), whose uncontrolled production has been associated with various inflammatory disease states as ...well as tumour proliferation and metastasis. Here we report the development and characterisation of an electrochemical peptide-based biosensor, which enables the detection of clinically relevant levels of HNE. The sensing platform was characterised in terms of its analytical performance, enzymatic cleavage kinetics and cross-reactivity and applied to the quantitative detection of protease activity from PMNs from human blood.
•We report the development of an electrochemical biosensor for human neutrophil elastase activity.•The sensing phase consisted in the immobilisation of a redox-labelled peptide onto a gold electrode via SAM.•The analytical characteristics were evaluated including limit of detection, reproducibility and selectivity.•The platform was applied for the detection of polymorphonuclear neutrophil (PMN) activity from human blood samples.
Mobile barnacle cypris larvae settle and metamorphose, transitioning to sessile juveniles with morphology and growth similar to that of adults. Because biofilms exist on immersed surfaces on which ...they attach, barnacles must interact with bacteria during initial attachment and subsequent growth. The objective of this study was to characterize the developing interface of the barnacle and substratum during this key developmental transition to inform potential mechanisms that promote attachment. The interface was characterized using confocal microscopy and fluorescent dyes to identify morphological and chemical changes to the interface and the status of bacteria present as a function of barnacle developmental stage. Staining revealed patchy material containing proteins and nucleic acids, reactive oxygen species amidst developing cuticle, and changes in bacteria viability at the developing interface. We found that as barnacles metamorphose from the cyprid to juvenile stage, proteinaceous materials with the appearance of coagulated liquid were released into and remained at the interface. It stained positive for proteins, including phosphoprotein, as well as nucleic acids. Regions of the developing cuticle and the patchy material itself stained for reactive oxygen species. Bacteria were absent until the cyprid was firmly attached, but populations died as barnacle development progressed. The oxidative environment may contribute to the cytotoxicity observed for bacteria and has the potential for oxidative crosslinking of cuticle and proteinaceous materials at the interface.
Electrochemical peptide-based biosensors are attracting significant attention for the detection and analysis of proteins. Here we report the optimisation and evaluation of an electrochemical ...biosensor for the detection of protease activity using self-assembled monolayers (SAMs) on gold surfaces, using trypsin as a model protease. The principle of detection was the specific proteolytic cleavage of redox-tagged peptides by trypsin, which causes the release of the redox reporter, resulting in a decrease of the peak current as measured by square wave voltammetry. A systematic enhancement of detection was achieved through optimisation of the properties of the redox-tagged peptide; this included for the first time a side-by-side study of the applicability of two of the most commonly applied redox reporters used for developing electrochemical biosensors, ferrocene and methylene blue, along with the effect of changing both the nature of the spacer and the composition of the SAM. Methylene blue-tagged peptides combined with a polyethylene-glycol (PEG) based spacer were shown to be the best platform for trypsin detection, leading to the highest fidelity signals (characterised by the highest sensitivity (signal gain) and a much more stable background than that registered when using ferrocene as a reporter). A ternary SAM (T-SAM) configuration, which included a PEG-based dithiol, minimised the non-specific adsorption of other proteins and was sensitive towards trypsin in the clinically relevant range, with a Limit of Detection (LoD) of 250pM. Kinetic analysis of the electrochemical response with time showed a good fit to a Michaelis–Menten surface cleavage model, enabling the extraction of values for kcat and KM. Fitting to this model enabled quantitative determination of the solution concentration of trypsin across the entire measurement range. Studies using an enzyme inhibitor and a range of real world possible interferents demonstrated a selective response to trypsin cleavage. This indicates that a PEG-based peptide, employing methylene blue as redox reporter, and deposited on an electrode as a ternary SAM configuration, is a suitable platform to develop clinically-relevant and quantitative electrochemical peptide-based protease biosensing.
•We report the optimisation of an electrochemical biosensor for protease activity.•A comparative study of the applicability of Fc and MB as redox reporters is presented.•MB-tagged peptides support enhanced electrochemical performance.•A T-SAM configuration using a PEG-based dithiol improved the analytical performance.
The growth of molluscan shell crystals is usually thought to be initiated from solution by extracellular organic matrix. We report a class of granulocytic hemocytes that may be directly involved in ...shell crystal production for oysters. On the basis of scanning electron microscopy (SEM) and x-ray microanalysis, these granulocytes contain calcium carbonate crystals, and they increase in abundance relative to other hemocytes following experimentally induced shell regeneration. Hemocytes are observed at the mineralization front using vital fluorescent staining and SEM. Some cells are observed releasing crystals that are subsequently remodeled, thereby at least augmenting matrix-mediated crystal-forming processes in this system.
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