28 results found
Belessiotis-Richards A, Larsen AH, Higgins SG, et al., 2022, Coarse-Grained Simulations Suggest Potential Competing Roles of Phosphoinositides and Amphipathic Helix Structures in Membrane Curvature Sensing of the AP180 N-Terminal Homology Domain., J Phys Chem B, Vol: 126, Pages: 2789-2797
The generation and sensing of membrane curvature by proteins has become of increasing interest to researchers with multiple mechanisms, from hydrophobic insertion to protein crowding, being identified. However, the role of charged lipids in the membrane curvature-sensing process is still far from understood. Many proteins involved in endocytosis bind phosphatidylinositol 4,5-bisphosphate (PIP2) lipids, allowing these proteins to accumulate at regions of local curvature. Here, using coarse-grained molecular dynamics simulations, we study the curvature-sensing behavior of the ANTH domain, a protein crucial for endocytosis. We selected three ANTH crystal structures containing either an intact, split, or truncated terminal amphipathic helix. On neutral membranes, the ANTH domain has innate curvature-sensing ability. In the presence of PIP2, however, only the domain with an intact helix senses curvature. Our work sheds light on the role of PIP2 and its modulation of membrane curvature sensing by proteins.
Belessiotis-Richards A, Larsen AH, Higgins SG, et al., 2022, Coarse-grained simulations suggest phosphoinositides and amphipathic helix structure play opposing roles in membrane curvature sensing of the AP180 N-terminal homology domain, The Journal of Physical Chemistry B: Biophysical Chemistry, Biomaterials, Liquids, and Soft Matter, ISSN: 1520-5207
The generation and sensing of membrane curvature by proteins has become of increasinginterest to researchers with multiple mechanisms, from hydrophobic insertion to protein crowding, beingidentified. However, the role of charged lipids in the membrane curvature sensing process is still far fromunderstood. Many proteins involved in endocytosis bind phosphatidylinositol 4,5-bisphosphate (PIP2)lipids, allowing these proteins to accumulate at regions of local curvature. Here, using coarse-grainedmolecular dynamics simulations, we study the curvature sensing behavior of the ANTH domain, a proteincrucial for endocytosis. We selected three ANTH crystal structures containing either an intact, split, ortruncated terminal amphipathic helix. On neutral membranes, the ANTH domain has innate curvaturesensing ability. In the presence of PIP2, however, only the domain with an intact helix senses curvature.Our work sheds light on the role of PIP2 and its modulation of membrane curvature sensing by proteins.
Bost JP, Ojansivu M, Munson MJ, et al., 2022, Novel endosomolytic compounds enable highly potent delivery of antisense oligonucleotides, Communications Biology, Vol: 5, ISSN: 2399-3642
The therapeutic and research potentials of oligonucleotides (ONs) have been hampered in part by their inability to effectively escape endosomal compartments to reach their cytosolic and nuclear targets. Splice-switching ONs (SSOs) can be used with endosomolytic small molecule compounds to increase functional delivery. So far, development of these compounds has been hindered by a lack of high-resolution methods that can correlate SSO trafficking with SSO activity. Here we present in-depth characterization of two novel endosomolytic compounds by using a combination of microscopic and functional assays with high spatiotemporal resolution. This system allows the visualization of SSO trafficking, evaluation of endosomal membrane rupture, and quantitates SSO functional activity on a protein level in the presence of endosomolytic compounds. We confirm that the leakage of SSO into the cytosol occurs in parallel with the physical engorgement of LAMP1-positive late endosomes and lysosomes. We conclude that the new compounds interfere with SSO trafficking to the LAMP1-positive endosomal compartments while inducing endosomal membrane rupture and concurrent ON escape into the cytosol. The efficacy of these compounds advocates their use as novel, potent, and quick-acting transfection reagents for antisense ONs.
Higgins S, Nogiwa Valdez A, Stevens M, 2022, Considerations for implementing electronic laboratory notebooks in an academic research environment, Nature Protocols, Vol: 17, Pages: 179-189, ISSN: 1750-2799
As research becomes predominantly digitalised, scientists have the option of usingelectronic laboratory notebooks to record and access entries. These systems can morereadily meet volume, complexity, accessibility and preservation requirements than papernotebooks. Whilst the technology can yield many benefits these can only be realised bychoosing a system that properly fulfils the requirements of a given context. This reviewexplores the factors that should be considered when introducing electronic laboratorynotebooks to an academically focused research group. We cite pertinent studies anddiscuss our own experience implementing a system within a multi-disciplinary researchenvironment. We also consider how the required financial and time investment is sharedbetween individuals and institutions. Finally, we discuss how electronic laboratory notebooksfit into the broader context of research data management. This article is not a productreview; it provides a framework for both the initial consideration of an electronic laboratorynotebook and the evaluation of specific software packages.
Brito L, Mylonaki I, Grigsby CL, et al., 2021, Genetic Enhancement of Epicardial Paracrine Signalling for Cardiac Regeneration, Publisher: LIPPINCOTT WILLIAMS & WILKINS, ISSN: 0009-7322
Becce M, Kloeckner A, Higgins S, et al., 2021, Assessing the impact of silicon nanowires on bacterial transformation and viability of Escherichia coli, Journal of Materials Chemistry B, Vol: 9, Pages: 4906-4914, ISSN: 2050-750X
We investigated the biomaterial interface between the bacteria Escherichia coli DH5α and silicon nanowire patterned surfaces. We optimised the engineering of silicon nanowire coated surfaces using metal-assisted chemical etching. Using a combination of focussed ion beam scanning electron microscopy, and cell viability and transformation assays, we found that with increasing interfacing force, cell viability decreases, as a result of increasing cell rupture. However, despite this aggressive interfacing regime, a proportion of the bacterial cell population remains viable. We found that the silicon nanowires neither resulted in complete loss of cell viability nor partial membrane disruption and corresponding DNA plasmid transformation. Critically, assay choice was observed to be important, as a reduction-based metabolic reagent was found to yield false-positive results on the silicon nanowire substrate. We discuss the implications of these results for the future design and assessment of bacteria–nanostructure interfacing experiments.
Wade J, Higgins SG, Heutz S, et al., 2021, In memoriam Alasdair James Campbell (11 May 1961-27 February 2021), Journal of Materials Chemistry C, Vol: 9, Pages: 6100-6102, ISSN: 2050-7526
Lin Y, Penna M, Spicer CD, et al., 2021, High-throughput peptide derivatization toward supramolecular diversification in microtiter plates, ACS Nano, Vol: 15, Pages: 4034-4044, ISSN: 1936-0851
The evolution of life on earth eventually leads to the emergence of species with increased complexity and diversity. Similarly, evolutionary chemical space exploration in the laboratory is a key step to pursue the structural and functional diversity of supramolecular systems. Here, we present a powerful tool that enables rapid peptide diversification and employ it to expand the chemical space for supramolecular functions. Central to this strategy is the exploitation of palladium-catalyzed Suzuki-Miyaura cross-coupling reactions to direct combinatorial synthesis of peptide arrays in microtiter plates under an open atmosphere. Taking advantage of this in situ library design, our results unambiguously deliver a fertile platform for creating a set of intriguing peptide functions including green fluorescent protein-like peptide emitters with chemically encoded emission colors, hierarchical self-assembly into nano-objects, and macroscopic hydrogels. This work also offers opportunities for quickly surveying the diversified peptide arrays and thereby identifying the structural factors that modulate peptide properties.
Belessiotis-Richards A, Higgins S, Sansom MSP, et al., 2020, Coarse Grained Simulations Suggest the Epsin N-Terminal Homology Domain Can Sense Membrane Curvature Without its Terminal Amphipathic Helix, ACS Nano, Vol: 14, Pages: 16919-6928, ISSN: 1936-0851
Nanoscale membrane curvature is a common feature in cell biology required for functions such as endocytosis, exocytosis and cell migration. These processes require the cytoskeleton to exert forces on the membrane to deform it. Cytosolic proteins contain specific motifs which bind to the membrane, connecting it to the internal cytoskeletal machinery. These motifs often bind charged phosphatidylinositol phosphate lipids present in the cell membrane which play significant roles in signaling. These lipids are important for membrane deforming processes, such as endocytosis, but much remains unknown about their role in the sensing of membrane nanocurvature by protein domains. Using coarse-grained molecular dynamics simulations, we investigated the interaction of a model curvature active protein domain, the epsin N-terminal homology domain (ENTH), with curved lipid membranes. The combination of anionic lipids (phosphatidylinositol 4,5-bisphosphate and phosphatidylserine) within the membrane, protein backbone flexibility, and structural changes within the domain were found to affect the domain's ability to sense, bind, and localize with nanoscale precision at curved membrane regions. The findings suggest that the ENTH domain can sense membrane curvature without the presence of its terminal amphipathic α helix <i>via</i> another structural region we have denoted as H3, re-emphasizing the critical relationship between nanoscale membrane curvature and protein function.
Higgins S, Lo Fiego A, Patrick I, et al., 2020, Organic bioelectronics: using highly conjugated polymers to interface with biomolecules, cells and tissues in the human body, Advanced Materials Technologies, Vol: 5, Pages: 1-35, ISSN: 2365-709X
Conjugated polymers exhibit interesting material and optoelectronic properties that makethem well-suited to the development of biointerfaces. Their biologically relevant mechanicalcharacteristics, ability to be chemically modified, and mixed electronic and ionic chargetransport are captured within the diverse field of organic bioelectronics. Conjugated polymershave been used in wide range of device architectures, and cell and tissue scaffolds. Thesedevices enable biosensing of many biomolecules, such as metabolites, nucleic acids and more.Devices can be used to both stimulate and sense the behavior of cells and tissues. Similarly,tissue interfaces permit interaction with complex organs, aiding both fundamental biologicalunderstanding and providing new opportunities for stimulating regenerative behaviors andbioelectronic based therapeutics. Applications of these materials are broad, and muchcontinues to be uncovered about their fundamental properties. This report covers the currentunderstanding of the fundamentals of conjugated polymer biointerfaces and their interactionswith biomolecules, cells and tissues in the human body. An overview of current materials anddevices is presented, along with highlighted major in vivo and in vitro applications. Finally,open research questions and opportunities are discussed.
Seong H, Higgins SG, Penders J, et al., 2020, Size-tunable nanoneedle arrays for influencing stem cell morphology, gene expression and nuclear membrane curvature, ACS Nano, Vol: 14, Pages: 5371-5381, ISSN: 1936-0851
High-aspect-ratio nanostructures have emerged as versatile platforms for intracellular sensing and biomolecule delivery. Here, we present a microfabrication approach in which a combination of reactive ion etching protocols was used to produce high-aspect-ratio, nondegradable silicon nanoneedle arrays with tip diameters that can be finely tuned between 20 and 700 nm. We used these arrays to guide the long-term culture of human mesenchymal stem cells (hMSCs). Notably, we used the nanoneedle tip diameter to control the morphology, nuclear size and F-actin alignment of interfaced hMSCs, and to regulate the expression of nuclear lamina genes, Yes-associated protein (YAP) target genes and focal adhesion genes. These topography-driven changes were attributed to signaling by Rho-family GTPase pathways, differences in the effective stiffness of the nanoneedle arrays and the degree of nuclear membrane impingement, with the latter clearly visualized using focused-ion beam scanning electron microscopy (FIB-SEM). Our approach to design high-aspect-ratio nanostructures will be broadly applicable to design biomaterials and biomedical devices used for long-term cell stimulation and monitoring.
Higgins S, Becce M, Belessiotis Richards A, et al., 2020, High-aspect-ratio nanostructured surfaces as biological metamaterials, Advanced Materials, Vol: 32, Pages: 1-44, ISSN: 0935-9648
Materials patterned with high-aspect-ratio nanostructures have features on similar lengthscales to cellular components. These surfaces are an extreme topography on the cellular leveland have become useful tools for perturbing and sensing the cellular environment. Motivationcomes from the ability of high-aspect-ratio nanostructures to deliver cargoes into cells andtissues, access the intracellular environment, and control cell behavior. These structuresdirectly perturb cells’ ability to sense and respond to external forces, influencing cell fate andenabling new mechanistic studies. Through careful design of their nanoscale structure, thesesystems act as biological metamaterials, eliciting unusual biological responses. Whilepredominantly used to interface eukaryotic cells, there is growing interest in non-animal andprokaryotic cell interfacing. Both experimental and theoretical studies have attempted todevelop a mechanistic understanding for the observed behaviors, predominantly focusing onthe cell – nanostructure interface. Here, we consider how high-aspect-ratio nanostructuredsurfaces are used to both stimulate and sense biological systems and discuss remainingresearch questions.
Higgins SG, Becce M, Seong H, et al., 2020, Nanoneedles and Nanostructured Surfaces for Studying Cell Interfacing, 7th International Conference on the Development of Biomedical Engineering, Publisher: SPRINGER-VERLAG SINGAPORE PTE LTD, Pages: 209-212, ISSN: 1680-0737
Higgins SG, 2019, Understanding scientists is key for science, NATURE MATERIALS, Vol: 18, Pages: 1144-1144, ISSN: 1476-1122
Belessiotis-Richards A, Higgins SG, Butterworth B, et al., 2019, Single-nanometer changes in nanopore geometry influence curvature, local properties, and protein localization in membrane simulations, Nano Letters, Vol: 19, Pages: 4770-4778, ISSN: 1530-6984
Nanoporous surfaces are used in many applications in intracellular sensing and drug delivery. However, the effects of such nanostructures on cell membrane properties are still far from understood. Here, we use coarse-grained molecular dynamics simulations to show that nanoporous substrates can stimulate membrane-curvature effects and that this curvature-sensing effect is much more sensitive than previously thought. We define a series of design parameters for inducing a nanoscale membrane curvature and show that nanopore taper plays a key role in membrane deformation, elucidating a previously unexplored fabrication parameter applicable to many nanostructured biomaterials. We report significant changes in the membrane area per lipid and thickness at regions of curvature. Finally, we demonstrate that regions of the nanopore-induced membrane curvature act as local hotspots for an increased bioactivity. We show spontaneous binding and localization of the epsin N-terminal homology (ENTH) domain to the regions of curvature. Understanding this interplay between the membrane curvature and nanoporosity at the biointerface helps both explain recent biological results and suggests a pathway for developing the next generation of cell-active substrates.
Mylonaki I, Amoli MS, Brito L, et al., 2019, Intracellular siRNA delivery using silicon nanoneedle arrays, British-Society-for-Gene-and-Cell-Therapy Autumn Conference, Publisher: MARY ANN LIEBERT, INC, Pages: A7-A7, ISSN: 1043-0342
Higgins S, 2018, Organising and running a STEM engagement stand inside a supermarket, Organising and running a STEM engagement stand inside a supermarket
Project report and evaluation for ‘Science in the Supermarket’
Higgins SG, Agostinelli T, Markham S, et al., 2017, Organic Diode Rectifiers Based on a High-Performance Conjugated Polymer for a Near-Field Energy-Harvesting Circuit, ADVANCED MATERIALS, Vol: 29, ISSN: 0935-9648
Higgins SG, Stevens MM, 2017, Extracting the contents of living cells, SCIENCE, Vol: 356, Pages: 379-380, ISSN: 0036-8075
Being able to monitor cells at different times is key to tracking fundamental cellular processes such as differentiation and cellular senescence, as well as disease progression and the effectiveness of drugs. However, most approaches are destructive and involve lysing the cells. Different time points can be studied by using parallel cell cultures, but the inferred changes could also be the result of cell heterogeneity (1, 2). Techniques for extracting small quantities of the cytosol for long-term tracking of a single cell's response must manipulate picoliter-scale volumes, maintain high cell viability, and give an accurate reflection of the cell's multiple biological components, as well as avoid influencing the ongoing development of the cell (see the figure) (1, 3). Cao et al. approached this problem by culturing cells on top of a random arrangement of hollow cylinders, which they call nanostraws (2). These 150-nm-diameter alumina tubes can sample 5 to 10% of proteins, messenger RNA (mRNA), and small molecules from the cells but only reduce cell viability by ∼5%. Their approach allows intracellular sampling and characterization at multiple time points from the same cells to track changes.
Higgins SG, Muir BVO, DellErba G, et al., 2016, Self-Aligned Organic Field-Effect Transistors on Plastic with Picofarad Overlap Capacitances and Megahertz Operating Frequencies, Applied Physics Letters, Vol: 108, ISSN: 0003-6951
Using a combination of nanoimprint lithography, gate-source/drain self-alignment, and gravure and inkjet printing, we fabricate organic field-effect transistors on flexible plastic substrates with gate-source and gate-drain electrode overlap capacitances of COL < 1 pF, equivalent to channel-width normalised capacitances of C*OL = 0.15–0.23 pF mm−1. We compare photopatterned and nanoimprint lithography patterned channels of L ≈ 3.8 μm and L ≈ 800 nm, respectively. The reduction in L was found on average to result in order of magnitude greater switching frequencies. Gravure printing the dielectric (versus photo-patterning) was found to yield an order of magnitude lower overlap capacitanceC*OL = 0.03 pF mm−1, at the expense of greater processing variation. Inkjet printed p- and n-type polymeric organic semiconductors were used to fabricate organic-field effect transistors with a peak cutoff frequencies of fS = 9.0 ± 0.3 MHz at VGS = 30 V, and transition frequencies of fT = 3.3 ± 0.2 MHz at VGS = 30 V.
Higgins S, Muir B, Dell'Erba G, et al., 2016, Complementary Organic Logic Gates on Plastic Formed by Self-Aligned Transistors with Gravure and Inkjet Printed Dielectric and Semiconductors, Advanced Electronic Materials, Vol: 2, ISSN: 2199-160X
Complementary organic field-effect transistors, inverters, NAND and NOR logic on plastic are demonstrated using a combination of nanoimprint lithography, self-alignment, gravure, and inkjet printing. Sub-micrometer channel lengths, electrode overlaps and sub-100 nm dielectrics are compared to photolithographically patterned equivalents, as are inkjet and gravure printed semiconductors.
Campbell AJ, Higgins SG, Muir BVO, et al., 2015, Indacenodithiophene–benzothiadiazole organic field-effect transistorswith gravure-printed semiconductor and dielectric on plastic, MRS Communications, Vol: 5, Pages: 599-603, ISSN: 2159-6867
We demonstrate the gravure printing of a high-performance indacenodithiophene (IDT) copolymer, indacenodithiophene–benzothiadiazole (C16IDT–BT), onto self-aligned organic field-effect transistor architectures on flexible plastic substrates. We observed that the combination of a gravure-printed dielectric with gravure-printed semiconductor yielded devices with higher mean-effective mobility than devices manufactured using photolithographically patterned dielectric. Peak mobilities of μ = 0.1 cm2 V−1 s−1 were measured, and exceed previous reports for non-printed C16IDT-BT on non-flexible silicon substrates.
Higgins SG, Muir BVO, Wade J, et al., 2015, Organic Transistors: Self-Aligned Megahertz Organic Transistors Solution-Processed on Plastic (Adv. Electron. Mater. 5/2015), Advanced Electronic Materials, Vol: 1
Higgins SG, Muir BVO, Wade J, et al., 2015, Self-aligned megahertz organic transistors solution-processed on plastic, Advanced Electronic Materials, Vol: 1, ISSN: 2199-160X
Higgins SG, Boughey FL, Hills R, et al., 2015, Quantitative Analysis and Optimization of Gravure Printed Metal Ink, Dielectric, and Organic Semiconductor Films, ACS APPLIED MATERIALS & INTERFACES, Vol: 7, Pages: 5045-5050, ISSN: 1944-8244
Ginzel R, Higgins SG, Mrowcynski P, et al., 2010, A deceleration system at the Heidelberg EBIT providing very slow highly charged ions for surface nanostructuring, NUCLEAR INSTRUMENTS & METHODS IN PHYSICS RESEARCH SECTION B-BEAM INTERACTIONS WITH MATERIALS AND ATOMS, Vol: 268, Pages: 2972-2976, ISSN: 0168-583X
Simon MC, Schwarz M, Epp SW, et al., 2010, Photoionization of N3+ and Ar8+ in an electron beam ion trap by synchrotron radiation, JOURNAL OF PHYSICS B-ATOMIC MOLECULAR AND OPTICAL PHYSICS, Vol: 43, ISSN: 0953-4075
Simon MC, Schwarz M, Schmitt BL, et al., 2009, Photoionization of ions in arbitrary charge states by synchrotron radiation in an electron beam ion trap, XXVI INTERNATIONAL CONFERENCE ON PHOTONIC, ELECTRONIC AND ATOMIC COLLISIONS, Vol: 194, ISSN: 1742-6588
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