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Journal articleWoubshete M, Cioccolo S, Byrne B, 2024,
Advances in membrane mimetic systems for manipulation and analysis of membrane proteins; detergents, polymers, lipids and scaffolds
, ChemPlusChem, Vol: 89, ISSN: 2192-6506Extracting membrane proteins from the hydrophobic environment of the biological membrane, in a physiologically relevant and stable state, suitable for downstream analysis remains a challenge. The traditional route to membrane protein extraction has been to use detergents and the last 15 years or so have seen a veritable explosion in the development of novel detergents with improved properties, making them more suitable for individual proteins and specific applications. There have also been significant advances in the development of encapsulation of membrane proteins in lipid based nanodiscs, either directly from the native membrane using polymers allowing effective capture of the protein and protein-associated membrane lipids, or via reconstitution of detergent extracted and purified protein into nanodiscs of defined lipid composition. All of these advances have been successfully applied to the study of membrane proteins via a range of techniques and there have been some spectacular membrane protein structures solved. In addition, the first detailed structural and biophysical analyses of membrane proteins retained within a biological membrane have been reported. Here we summarise and review the recent advances with respect to these new agents and systems for membrane protein extraction, reconstitution and analysis.
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Journal articleLai H, Kolanko M, Li LM, et al., 2024,
Population incidence and associated mortality of urinary tract infection in people living with dementia
, JOURNAL OF INFECTION, Vol: 88, ISSN: 0163-4453 -
Journal articleLu Y, Duman R, Beilsten-Edmands J, et al., 2024,
Ray-tracing analytical absorption correction for X-ray crystallography based on tomographic reconstructions
, JOURNAL OF APPLIED CRYSTALLOGRAPHY, Vol: 57, Pages: 649-658, ISSN: 1600-5767 -
Journal articleGallo A, Mansueto S, Emendato A, et al., 2024,
α-Synuclein and Mitochondria: Probing the Dynamics of Disordered Membrane-protein Regions Using Solid-State Nuclear Magnetic Resonance
, JACS AU -
Journal articleGosiamemang T, Heng JYY, 2024,
Simple one-pot fabrication of durable superhydrophobic cotton wool using octadecyltrimethoxysilane modified silica-sol for oil-water separation
, Separation and Purification Technology, Vol: 336, ISSN: 0950-4214A fluorine-free superhydrophobic cotton is successfully fabricated using a simple and cost-effective one-pot synthesis method, employing octadecyltrimethoxysilane (ODTMS) as a modifying silane in a silica-sol. Unlike conventional techniques that utilise coupling agents or post-synthesis curing methods to enhance coating stability, this approach eliminates the need for extra chemicals thus saving time and reducing fabrication cost. The prepared cotton demonstrates remarkable superhydrophobicity, as evidenced by a water contact angle of 159°. Additionally, it exhibits exceptional durability and stability when exposed to harsh acidic and alkaline solutions, as well as during laundering test. Moreover, the cotton displays excellent oil–water separation efficiency (>99.0 %) and maintains consistent performance throughout 20 reuse cycles, highlighting its high reusability. Furthermore, the modified cotton is highly effective for separating oil-in-water emulsions, even for highly stable emulsions, achieving separation efficiencies above 90.0 %. The fabrication process is simple and environmentally safe, offering an economical and viable solution for industries handling oily wastewater. Overall, this fluorine-free superhydrophobic cotton presents a promising and cost-effective option for various industrial applications.
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Journal articleMaertens GN, Purcell DFJ, Rosadas C, et al., 2024,
Why not eliminate HTLV-1 while eliminating HIV-1?
, LANCET, Vol: 403, Pages: 2288-2289, ISSN: 0140-6736 -
Journal articleYuen ELH, Leary AY, Clavel M, et al., 2024,
A RabGAP negatively regulates plant autophagy and immune trafficking
, CURRENT BIOLOGY, Vol: 34, ISSN: 0960-9822 -
Journal articleFecht S, Paracuellos P, Subramoni S, et al., 2024,
Functionality of chimeric TssA proteins in the type VI secretion system reveals sheath docking specificity within their N-terminal domains
, Nature Communications, Vol: 15, ISSN: 2041-1723The genome of Pseudomonas aeruginosa encodes three type VI secretion systems, each comprising a dozen distinct proteins, which deliver toxins upon T6SS sheath contraction. The least conserved T6SS component, TssA, has variations in size which influence domain organisation and structure. Here we show that the TssA Nt1 domain interacts directly with the sheath in a specific manner, while the C-terminus is essential for oligomerisation. We built chimeric TssA proteins by swapping C-termini and showed that these can be functional even when made of domains from different TssA sub-groups. Functional specificity requires the Nt1 domain, while the origin of the C-terminal domain is more permissive for T6SS function. We identify two regions in short TssA proteins, loop and hairpin, that contribute to sheath binding. We propose a docking mechanism of TssA proteins with the sheath, and a model for how sheath assembly is coordinated by TssA proteins from this position.
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Journal articleLiang E, Verma V, Guo M, et al., 2024,
Sequence-dependent dipeptide solubility in ethanol-water and DMSO-water solutions
, Journal of Molecular Liquids, Vol: 402, ISSN: 0167-7322Reliable solubility data is crucial for controlling and optimising crystallisation conditions, to provide a cost-effective purification method with the potential for scaling up. This study investigated the effect of amino acid sequence on determining peptides solubility behaviours. Four dipeptides with different sequences and combinations of alanine and glycine were selected to study their temperature dependent (283.15 K to 313.15 K) aqueous solubilities and solubility in the presence of ethanol and DMSO as antisolvent at 298.15 K. Aqueous solubility data of dipeptides presents the following order: glycyl-L-alanine (gly-ala) > L-alanyl-L-alanine (ala-ala) > glycyl glycine (gly-gly) > L-alanyl glycine (ala-gly). This solubility order has been substantiated by solvation free energy calculations using molecular dynamics (MD) simulations, providing insights into the underlying molecular interactions that govern these solubility patterns. The solubility data also demonstrated difference in the temperature dependency that: gly-gly > gly-ala > ala-gly > ala-ala, corresponding to their dissolution enthalpy order. In the presence of antisolvent range from 0 % to 80 % (by mass) concentration, peptide’s solubility was more susceptible to DMSO than ethanol. Accurate prediction of the solubilities validated the effectiveness of non-random two-liquid (NRTL) model for peptide studies.
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ReportFreemont P, Chang MW, Hook-Barnard I, et al., 2024,
Engineering biology metrics and technical standards for the global bioeconomy
, Engineering biology metrics and technical standards for the global bioeconomy -
Conference paperIbrahim T, Vuolo C, Nayyar BG, et al., 2024,
AI guided discovery of cross kingdom autophagy modulators during host-pathogen interactions
, 14th Congress on International-Society-for-Molecular-Plant-Microbe-Interactions (IS-MPMI), Publisher: AMER PHYTOPATHOLOGICAL SOC, Pages: 141-141, ISSN: 0894-0282 -
Journal articleRostøl JT, Quiles-Puchalt N, Iturbe-Sanz P, et al., 2024,
Bacteriophages avoid autoimmunity from cognate immune systems as an intrinsic part of their life cycles
, Nature Microbiology, Vol: 9, Pages: 1312-1324, ISSN: 2058-5276Dormant prophages protect lysogenic cells by expressing diverse immune systems, which must avoid targeting their cognate prophages upon activation. Here we report that multiple Staphylococcus aureus prophages encode Tha (tail-activated, HEPN (higher eukaryotes and prokaryotes nucleotide-binding) domain-containing anti-phage system), a defence system activated by structural tail proteins of incoming phages. We demonstrate the function of two Tha systems, Tha-1 and Tha-2, activated by distinct tail proteins. Interestingly, Tha systems can also block reproduction of the induced tha-positive prophages. To prevent autoimmunity after prophage induction, these systems are inhibited by the product of a small overlapping antisense gene previously believed to encode an excisionase. This genetic organization, conserved in S. aureus prophages, allows Tha systems to protect prophages and their bacterial hosts against phage predation and to be turned off during prophage induction, balancing immunity and autoimmunity. Our results show that the fine regulation of these processes is essential for the correct development of prophages' life cycle.
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Journal articleGupta SS, Maetzig T, Maertens GN, et al., 2024,
Correction for Gupta et al., “Bromo- and extraterminal domain chromatin regulators serve as cofactors for murine leukemia virus integration”
, Journal of Virology, Vol: 98, ISSN: 0022-538X -
Journal articleChen SW, Barritt JD, Cascella R, et al., 2024,
Structure-Toxicity Relationship in Intermediate Fibrils from α-Synuclein Condensates
, JOURNAL OF THE AMERICAN CHEMICAL SOCIETY, Vol: 146, Pages: 10537-10549, ISSN: 0002-7863 -
Journal articleCrone MA, Hakki S, Fenn J, et al., 2024,
Rapid emergence of transmissible SARS-CoV-2 variants in mild community cases
, Microbiology Spectrum, Vol: 12, ISSN: 2165-0497 -
Journal articleKabasakal BV, Cotton CAR, Murray JW, 2024,
Dynamic lid domain of Chloroflexus aurantiacus Malonyl-CoA reductase controls the reaction.
, Biochimie, Vol: 219, Pages: 12-20, ISSN: 0300-9084Malonyl-Coenzyme A Reductase (MCR) in Chloroflexus aurantiacus, a characteristic enzyme of the 3-hydroxypropionate (3-HP) cycle, catalyses the reduction of malonyl-CoA to 3-HP. MCR is a bi-functional enzyme; in the first step, malonyl-CoA is reduced to the free intermediate malonate semialdehyde by the C-terminal region of MCR, and this is further reduced to 3-HP by the N-terminal region of MCR. Here we present the crystal structures of both N-terminal and C-terminal regions of the MCR from C. aurantiacus. A catalytic mechanism is suggested by ligand and substrate bound structures, and structural and kinetic studies of MCR variants. Both MCR structures reveal one catalytic, and one non-catalytic SDR (short chain dehydrogenase/reductase) domain. C-terminal MCR has a lid domain which undergoes a conformational change and controls the reaction. In the proposed mechanism of the C-terminal MCR, the conversion of malonyl-CoA to malonate semialdehyde is based on the reduction of malonyl-CoA by NADPH, followed by the decomposition of the hemithioacetal to produce malonate semialdehyde and coenzyme A. Conserved arginines, Arg734 and Arg773 are proposed to play key roles in the mechanism and conserved Ser719, and Tyr737 are other essential residues forming an oxyanion hole for the substrate intermediates.
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Journal articleGriffiths D, Anderson M, Richardson K, et al., 2024,
Cyclic Ion Mobility for Hydrogen/Deuterium Exchange-Mass Spectrometry Applications
, ANALYTICAL CHEMISTRY, ISSN: 0003-2700 -
Journal articleBaquero F, Beis K, Craik DJ, et al., 2024,
The pearl jubilee of microcin J25: thirty years of research on an exceptional lasso peptide
, NATURAL PRODUCT REPORTS, Vol: 41, Pages: 469-511, ISSN: 0265-0568 -
Journal articlevan Haaren C, Byrne B, Kazarian SG, 2024,
Study of monoclonal antibody aggregation at the air-liquid interface under flow by ATR-FTIR spectroscopic imaging
, Langmuir: the ACS journal of surfaces and colloids, Vol: 40, Pages: 5858-5868, ISSN: 0743-7463Throughout bioprocessing, transportation, and storage, therapeutic monoclonal antibodies (mAbs) experience stress conditions that may cause protein unfolding and/or chemical modifications. Such structural changes may lead to the formation of aggregates, which reduce mAb potency and may cause harmful immunogenic responses in patients. Therefore, aggregates need to be detected and removed or ideally prevented from forming. Air-liquid interfaces, which arise during various stages of bioprocessing, are one of the stress factors causing mAb aggregation. In this study, the behavior of an immunoglobulin G (IgG) at the air-liquid interface was investigated under flow using macro attenuated total reflection Fourier transform infrared (ATR-FTIR) spectroscopic imaging. This chemically specific imaging technique allows observation of adsorption of IgG to the air-liquid interface and detection of associated secondary structural changes. Chemical images revealed that IgG rapidly accumulated around an injected air bubble under flow at 45 °C; however, no such increase was observed at 25 °C. Analysis of the second derivative spectra of IgG at the air-liquid interface revealed changes in the protein secondary structure associated with increased intermolecular β-sheet content, indicative of aggregated IgG. The addition of 0.01% w/v polysorbate 80 (PS80) reduced the amount of IgG at the air-liquid interface in a static setup at 30 °C; however, this protective effect was lost at 45 °C. These results suggest that the presence of air-liquid interfaces under flow may be detrimental to mAb stability at elevated temperatures and demonstrate the power of ATR-FTIR spectroscopic imaging for studying the structural integrity of mAbs under bioprocessing conditions.
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Journal articlePerrett S, Chatrchyan V, Buckup T, et al., 2024,
Application of density matrix Wigner transforms for ultrafast macromolecular and chemical x-ray crystallography
, JOURNAL OF CHEMICAL PHYSICS, Vol: 160, ISSN: 0021-9606 -
Journal articleSmith HE, Mackenzie AM, Seddon C, et al., 2024,
The use of NADH anisotropy to investigate mitochondrial cristae alignment
, SCIENTIFIC REPORTS, Vol: 14, ISSN: 2045-2322 -
Journal articleGardner S, Jin Y, Fyfe PK, et al., 2024,
Structural insights into IL-11-mediated signalling and human IL6ST variant-associated immunodeficiency
, Nature Communications, Vol: 15, ISSN: 2041-1723IL-11 and IL-6 activate signalling via assembly the cell surface receptor gp130; however, it is unclear how signals are transmitted across the membrane to instruct cellular responses. Here we solve the cryoEM structure of the IL-11 receptor recognition complex to discover how differences in gp130-binding interfaces may drive signalling outcomes. We explore how mutations in the IL6ST gene encoding for gp130, which cause severe immune deficiencies in humans, impair signalling without blocking cytokine binding. We use cryoEM to solve structures of both IL-11 and IL-6 complexes with a mutant form of gp130 associated with human disease. Together with molecular dynamics simulations, we show that the disease-associated variant led to an increase in flexibility including motion within the cytokine-binding core and increased distance between extracellular domains.However, these distances are minimized as the transmembrane helix exits the membrane, suggesting a stringency in geometry for signalling and dimmer switch mode of action.
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Journal articleChukhutsina VU, Hutchison CDM, van Thor JJ, 2024,
The Carbonyl Group in b2 of the Carotenoid Tunes the Photocycle Kinetics in Orange Carotenoid Protein
, JOURNAL OF MOLECULAR BIOLOGY, Vol: 436, ISSN: 0022-2836 -
Journal articleMazza T, Roumeliotis TI, Garitta E, et al., 2024,
Structural basis for the modulation of MRP2 activity by phosphorylation and drugs
, Nature Communications, Vol: 15, ISSN: 2041-1723Multidrug resistance-associated protein 2 (MRP2/ABCC2) is a polyspecific efflux transporter of organic anions expressed in hepatocyte canalicular membranes. MRP2 dysfunction, in Dubin-Johnson syndrome or by off-target inhibition, for example by the uricosuric drug probenecid, elevates circulating bilirubin glucuronide and is a cause of jaundice. Here, we determine the cryo-EM structure of rat Mrp2 (rMrp2) in an autoinhibited state and in complex with probenecid. The autoinhibited state exhibits an unusual conformation for this class of transporter in which the regulatory domain is folded within the transmembrane domain cavity. In vitro phosphorylation, mass spectrometry and transport assays show that phosphorylation of the regulatory domain relieves this autoinhibition and enhances rMrp2 transport activity. The in vitro data is confirmed in human hepatocyte-like cells, in which inhibition of endogenous kinases also reduces human MRP2 transport activity. The drug-bound state reveals two probenecid binding sites that suggest a dynamic interplay with autoinhibition. Mapping of the Dubin-Johnson mutations onto the rodent structure indicates that many may interfere with the transition between conformational states.
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Journal articlevan Thor J, 2024,
Kilohertz droplet-on-demand serial femtosecond crystallography at the European XFEL station FXE
, Structural Dynamics, Vol: 11, ISSN: 2329-7778X-ray Free Electron Lasers (XFELs) allow the collection of high-quality serial femtosecond crystallography data. The next generation of megahertz superconducting FELs promises to drastically reduce data collection times, enabling the capture of more structures with higher signal-to-noise ratios and facilitating more complex experiments. Currently, gas dynamic virtual nozzles (GDVNs) stand as the sole delivery method capable of best utilizing the repetition rate of megahertz sources for crystallography. However, their substantial sample consumption renders their use impractical for many protein targets in serial crystallography experiments. Here, we present a novel application of a droplet-on-demand injection method, which allowed operation at 47 kHz at the European XFEL (EuXFEL) by tailoring a multi-droplet injection scheme for each macro-pulse. We demonstrate a collection rate of 150 000 indexed patterns per hour. We show that the performance and effective data collection rate are comparable to GDVN, with a sample consumption reduction of two orders of magnitude. We present lysozyme crystallographic data using the Large Pixel Detector at the femtosecond x-ray experiment endstation. Significant improvement of the crystallographic statistics was made by correcting for a systematic drift of the photon energy in the EuXFEL macro-pulse train, which was characterized from indexing the individual frames in the pulse train. This is the highest resolution protein structure collected and reported at the EuXFEL at 1.38 Å resolution.
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Journal articleYoon S, Bae HE, Hariharan P, et al., 2024,
Rational approach to improve detergent efficacy for membrane protein stabilization
, Bioconjugate Chemistry, Vol: 35, Pages: 223-231, ISSN: 1043-1802Membrane protein structures are essential for the molecular understanding of diverse cellular processes and drug discovery. Detergents are not only widely used to extract membrane proteins from membranes but also utilized to preserve native protein structures in aqueous solution. However, micelles formed by conventional detergents are suboptimal for membrane protein stabilization, necessitating the development of novel amphiphilic molecules with enhanced protein stabilization efficacy. In this study, we prepared two sets of tandem malonate-derived glucoside (TMG) variants, both of which were designed to increase the alkyl chain density in micelle interiors. The alkyl chain density was modulated either by reducing the spacer length (TMG-Ms) or by introducing an additional alkyl chain between the two alkyl chains of the original TMGs (TMG-Ps). When evaluated with a few membrane proteins including a G protein-coupled receptor, TMG-P10,8 was found to be substantially more efficient at extracting membrane proteins and also effective at preserving protein integrity in the long term compared to the previously described TMG-A13. This result reveals that inserting an additional alkyl chain between the two existing alkyl chains is an effective way to optimize detergent properties for membrane protein study. This new biochemical tool and the design principle described have the potential to facilitate membrane protein structure determination.
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Thesis dissertationMorrison A, 2024,
Development of Improved Cultivation Methods for Environmental Microorganisms
Cultivation of bacteria remains an essential prerequisite for numerous research and biotechnological applications. Despite their ubiquity, only a minority of environmental bacteria are cultivable using standard techniques. Therefore, a vast microbial ‘dark matter’ awaits exploration for valuable therapeutics or research potential. Despite innovations in enhanced cultivation techniques, limitations remain including throughput, dependencies on environmental factors or abrupt transitions of microorganisms from native to laboratory conditions. This thesis addresses these challenges with the development, optimisation, and experimental assessment of two novel cultivation methodologies. The first methodology allows for the gradual transition and acclimatisation of microorganisms from native environments to culture media using the novel Enhanced Domestication (EDEN) device. Compared to the instantaneous transition, acclimatisation of pond water bacteria to R2A culture media significantly enhanced cultivation diversity, colony yield and greater taxonomic range of isolates including those previously reported to be recalcitrant to cultivation. Moreover, likely novel taxa cultivated with EDEN exhibited antimicrobial activity against methicillin-resistant Staphylococcus aureus. The second methodology involves the development of a cell encapsulation apparatus, termed Bacterial Encapsulation and Containment (BEAD), enabling microinjection and cultivation of bacteria within alginate hydrogel beads using an automated pneumatic pump system. The performance of in situ cultivation with BEAD-encapsulated bacteria were assessed using the newly developed cultivation cassettes growth chambers. The findings propose enhanced cultivation strategies to unlock the biosynthetic potential of uncultured environmental bacteria.
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Journal articleChadha D, Heng J, 2024,
A scoping review of professional skills development in engineering education from 1980-2020
, Cogent Education, Vol: 14, ISSN: 2331-186XEngineering students are expected to develop their professional skills throughout the course of their degrees. Yet, there is no clear consensus among educators of which skills are being developed or how this is done effectively. This scoping review has been produced to draw together and disseminate information on effective techniques for developing six specific professional skills (communication, teamwork, problem solving, leadership, project management and entrepreneurship) and tracing the evolution of these skills in engineering education. Using search engines and backwards and forwards snowballing, a scoping literature review on skills development in engineering was conducted, encompassing a 40-year time span (1980–2020). Specified search terms, and inclusion criteria were applied, and findings have been presented quantitatively and qualitatively. In total, 165 studies were included. Key findings indicate skills are more nuanced and accepted today than they were in 1980, with an increased focus on specific skill areas. Additionally, more sophisticated integrated delivery methods are used to teach skills, although short courses remain popular. By synthesising data in this way, professional educators can identify what skill areas to focus on, and how to teach these skills for effective development among students. Gaps in knowledge are also identified through this review. This study is significant in that it has created an exceptionally useful roadmap of skills development, which is of benefit to the engineering community and beyond.
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Journal articleVieira MFM, Hernandez G, Zhong Q, et al., 2024,
The pathogen-encoded signalling receptor Tir exploits host-like intrinsic disorder for infection
, COMMUNICATIONS BIOLOGY, Vol: 7 -
Conference paperGirvan P, Jalal ASB, McCormack EA, et al., 2024,
Nucleosome flipping drives selectivity and processivity of histone exchange by SWR1 complex
, Publisher: CELL PRESS, Pages: 499A-500A, ISSN: 0006-3495
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