I22-Small angle scattering & Diffraction
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Tayyaba
Rabnawaz
,
Nathanael
Leung
,
Leonard C.
Nielsen
,
Robert A.
Harper
,
Richard M.
Shelton
,
Gabriel
Landini
,
Tim
Snow
,
Andy
Smith
,
Nick
Terrill
,
Marianne
Liebi
,
Tan
Sui
Diamond Proposal Number(s):
[20285]
Abstract: Dental caries, one of the most prevalent non-communicable diseases worldwide, is characterised by the progressive deterioration of the structure and mechanical properties of dental hard tissues. In human teeth, dentine is the most abundant mineralised tissue, forming the primary support material. To assess changes in the mechanical properties of dentine caused by dental caries and acid erosion, it is crucial to understand the relationship between organic and inorganic dentine components and their organisation into a 3D anisotropic structure at the nanoscale. Over the past 20 years, alterations in dentine structure caused by caries and artificial demineralisation have been reported using conventional microscopy techniques. However, due to the limited spatial resolution of these techniques, the 3D structural organisation including orientation and degree of alignment of mineralised collagen fibrils at the nanoscale, has not been fully explored. This study investigated alterations in the 3D structure of normal, carious and artificially demineralised dentine using SAXS tensor tomography (SASTT). This technique enabled the observation of differences in the local orientation of organic and inorganic components, as well as variations in local scattering intensity, resulting from natural caries and artificial demineralisation. In comparison to normal dentine, caries caused minor orientational differences of both components but had a major impact on the local X-ray scattering intensity. After artificial demineralisation of the dentine, most of the mineral was lost in the outer layers, resulting in a greater reduction in scattering intensity than that caused by caries.
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Mar 2026
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B21-High Throughput SAXS
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Diamond Proposal Number(s):
[29895]
Open Access
Abstract: A lipopeptide is designed that contains an epitope from simian virus T-antigen (SV40T, PKKKRKV) conjugated to an N-terminal palmitoyl (C16-) moiety, with the aim to act as an effective cell-penetrating lipopeptide, with additional aggregation propensity conferred by the lipid chain. A combination of cryo-TEM and small-angle X-ray scattering (SAXS) is used to show that the lipopeptide forms micelles, but mixtures with DNA lead to formation of fractal cluster-like co-assemblies due to intercalation of the DNA and peptide. Spectroscopic studies using fluorescence and circular dichroism (along with fiber X-ray diffraction) show that the peptide interacts with DNA and inserts into the groove. Confocal microscopy along with flow cytometry confirms delivery of DNA into both HeLa and mouse embryonic stem cells (mESCs) in pluripotent state, and the system shows excellent cytocompatibility as confirmed by MTT assays. Our data indicate that the lipopeptide may outperform the DNA transfection agent lipofectamine in DNA delivery into these stem cells and it enables DNA delivery into the cytoplasm and nucleus.
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Mar 2026
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VMXi-Versatile Macromolecular Crystallography in situ
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Diamond Proposal Number(s):
[28534, 34263]
Open Access
Abstract: Schistosoma mansoni cathepsin D1 (SmCD1) has been shown to be an essential enzyme for helminth metabolism due to its role in haemoglobin degradation: a key amino-acid source for the developing parasite. Therefore, the enzyme is a potential target for the development of antischistosomal inhibitors. SmCD1 has significant sequence identity to cathepsin D-like proteases found in other schistosome species and homology to mammalian aspartic proteases. Here, we report the first crystal structures of a helminth cathepsin D, SmCD1, and have identified a single-domain antibody (nanobody) that specifically binds to SmCD1 with nanomolar affinity but does not recognize human cathepsin D. We have mapped the epitope of the nanobody by determining the crystal structure of the enzyme–nanobody complex, revealing the conformation of SmCD1 in the propeptide-bound state.
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Feb 2026
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DIAD-Dual Imaging and Diffraction Beamline
I14-Hard X-ray Nanoprobe
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Abstract: High-resolution characterisation of biomaterials across multiple length scales to investigate their effect on (re)mineralisation can inform the development of effective interventions for clinical conditions such as dental caries, a disease with an estimated global economic burden of approximately $245 billion. This thesis presents a multi-modal, synchrotron-based approach on novel instruments to study the role of self-assembling peptide P11-4 in dental enamel (re)mineralisation, with the aim of elucidating its mechanism of action for potential optimisation to treat early caries lesions non-invasively, and to use P11-4 as a model system for the development of a liquid flow cell that can be used to characterise biomimetic materials in situ using synchrotron X-ray diffraction (XRD) and X-ray microtomography (XMT).
X-ray fluorescence combined with differential phase contrast imaging on the I14 beamline, together with XRD and XMT on the Dual Imaging and Diffraction beamline at Diamond Light Source, along with complementary laboratory-based techniques, were employed to characterise P11-4.
P11-4 accelerated the initial kinetics of the mineralisation process compared to the control, via the provision of calcium-binding sites, and controlled the mineral deposition process, mimicking the role of enamel matrix proteins during biomineralisation. The chemical model used for artificial demineralisation to create caries-like lesions resulted in preferential demineralisation of the enamel prisms. Within the caries-like lesions, the developed flow cell demonstrated that P11-4 promoted deep remineralisation of the lesion via the gradual formation of organised apatite structures within one specific population of crystallites, likely corresponding to the prisms. The organisation of crystallites within the regenerated structure is comparable to healthy enamel, highlighting its role in restoring the organised structure lost due to caries, and its significance as a non-invasive clinical treatment.
The methodology presented in this thesis can be applied to analyse lesions and characterise other biomaterials/proteins, and the flow cell is available to other users.
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Feb 2026
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I04-Macromolecular Crystallography
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Diamond Proposal Number(s):
[31440, 37593]
Open Access
Abstract: Angiotensin I-converting enzyme (ACE) is a zinc-dependent dipeptidyl carboxypeptidase involved in blood pressure regulation through proteolysis of angiotensin I (Ang-I) into the potent vasoconstrictor, angiotensin II (Ang-II). Inhibition of ACE is therefore used for the treatment of hypertension, heart failure, myocardial infarction, stroke and chronic kidney disease. Current ACE inhibitors (ACEi) bind both the N- and C-catalytic domains of ACE (referred to as nACE and cACE), and this has been linked to the occurrence of side effects due to the wide substrate specificity of ACE. The development of domain selective ACEi with reduced side effects is therefore key for improved therapeutic intervention. Understanding how current ACEi bind nACE and cACE, and their differences in domain selectivity should aid structure-based development of more selective ACEi by identifying different chemical groups that increase or decrease selectivity. We present the kinetic and structural characterisation of nACE and cACE with three thiolate ACEi, captopril (Ki, nACE = 2.53 nm and cACE = 2.04 nm), rentiapril (monomer Ki, nACE = 2.22 nm and cACE = 6.77 nm) and zofenoprilat (Ki, nACE = 2.86 nm and cACE = 0.61 nm). Detailed structural analysis indicated that the S2′ subsite likely contributes to the variation in domain selectivity observed for rentiapril and zofenoprilat due to differences in hydrophobicity and displacement of water molecules that contribute to ACE's hydration shell. Interestingly, in the cACE crystal structure, rentiapril bound as a dimer, and kinetic data revealed that both the monomeric and dimeric (dimer Ki, nACE = 15.11 nm and cACE = 36.38 nm) forms of rentiapril inhibit ACE with nanomolar affinity.
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Feb 2026
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B21-High Throughput SAXS
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Diamond Proposal Number(s):
[29895]
Abstract: A hydrogel formed by a short peptide is presented that exhibits remarkable stimuli-responsiveness and plasticity, undergoing a morphological transformation from nanofibers to nanospheres in the presence of monovalent (Li+, Na+, K+) or trivalent (Al3+, Fe3+) metal ions under physiological conditions. The nanofibrillar structure of the hydrogel was examined using transmission electron microscopy (TEM), atomic force microscopy (AFM), small-angle X-ray scattering (SAXS), and X-ray diffraction (XRD) studies and atomistic molecular dynamics simulations, in complement, to explain the nanostructural transitions at the microscopic level. Interestingly, exposure to divalent metal ions (Mg2+, Ca2+, Co2+, Ni2+) induces a unique shrinking (syneresis) behavior, accompanied by a morphological shift to nanoribbons. Both simulations and SAXS analysis confirm that these ions cause a contraction in the packing of gelator peptides, significantly reducing the interpeptide distance. This ion-specific adaptability confers tunable physicochemical properties and morphological plasticity. Hydrogels incorporating mono- or trivalent ions exhibit enhanced thermal stability and mechanical strength relative to ion-free counterparts, underscoring the reinforcing role of metal coordination. Strikingly, shrunken gels formed in the presence of divalent ions display even greater stiffness than freshly prepared gels in the absence of any metal ions, suggesting that syneresis acts as a postassembly strengthening mechanism. These findings highlight a versatile, stimuli-responsive soft material in which ion-peptide interactions orchestrate nanoscale morphology, mesoscale network architecture, and macroscopic mechanical performance-opening avenues for adaptive hydrogel systems in targeted biomedical, sensing, and controlled-release applications.
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Feb 2026
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Open Access
Abstract: Perspectives surrounding Cannabis use have transformed over the past decade. This shift in perspective has been noted in pregnant populations in Canada and the US, where various investigations report that the use of Cannabis in pregnancy is increasingly commonplace. There is some evidence indicating that Δ9-tetrahydrocannabinol (THC), the main intoxicating phytocannabinoid found within Cannabis flower, may influence the biochemical composition of lipids within the developing fetal brain. The aim of this study was to apply multimodal biospectroscopic imaging techniques, X-ray fluorescence imaging (XFI) and Fourier transform mid-infrared spectromicroscopy (FTIR), to investigate the biochemical and biomolecular changes underlying the distinct behavioral phenotypes identified previously. XFI was used to investigate the presence of metal, nonmetal, and alkali dysregulation, while FTIR provided information on neurochemical dysbiosis within the brains of offspring exposed to THC (3 mg/kg; i.p.) or vehicle (VEH). The THC offspring exhibited decreased copper (Cu) concentrations within the perimeter of their corpus callosum, as identified by XFI. FTIR hyperspectral data from the brain revealed noteworthy changes in peaks associated with lipid methylene (CH2as), carbohydrates, and peak ratios identifying changes in the lipid structure and the relative content of lipids, cholesterol esters, and cholesterols to saturated fatty acids. These changes were particularly evident in the hippocampus, where THC offspring exhibited increased CH2as, lipid esters, phosphate, protein, and unsaturation levels of lipids. The biochemical changes seen in the FTIR spectra were modest, with THC offspring showing an increase in the number of structural changes of lipids in the corpus callosum and an increase in protein in the lateral ventricle. This study supports the usefulness of these techniques to detect subtle changes in biomolecular composition within brain tissues exposed to gestational THC. These results contribute to the growing body of knowledge unraveling the complex effects of THC on fetal neurodevelopmental trajectories.
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Feb 2026
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I04-Macromolecular Crystallography
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Qiangqiang
Wei
,
Ashley J.
Taylor
,
Nagaraju
Miriyala
,
Mahesh A.
Barmade
,
Zachary O.
Gentry
,
Jordan
Anderson-Daniels
,
Kevin B.
Teuscher
,
Mackenzie M.
Crow
,
Chideraa
Apakama
,
Taylor M.
South
,
Tyson A.
Rietz
,
Kangsa
Amporndanai
,
Jason
Phan
,
John L.
Sensintaffar
,
Mark
Denison
,
Taekyu
Lee
,
Stephen W.
Fesik
Open Access
Abstract: The papain-like protease (PLPro) plays a key role in SARS-CoV-2 replication and represents a promising target for the development of new antiviral therapies. Previous efforts to develop fragment-derived inhibitors of PLPro led to the identification of a novel class of spiro[chromane-2,4′-piperidin]-4-one inhibitors exemplified by lead compound 7. High-resolution covalent cocrystal structures and molecular dynamics simulations were utilized to guide the development of a series of low-nanomolar irreversible PLPro inhibitors, with lead compound 45 demonstrating strong enzymatic inhibition (IC50 = 0.059 μM at T = 60 min) and antiviral activity in A549 cells (EC50 = 2.1 μM at 48 hpi). This novel class of inhibitors represents a promising avenue for the development of therapeutics to overcome the potential of drug-resistant viral strains and future coronavirus outbreaks.
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Feb 2026
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I20-Scanning-X-ray spectroscopy (XAS/XES)
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Diamond Proposal Number(s):
[31906, 39961]
Open Access
Abstract: Mercury (Hg) is a global environmental concern due to its microbial conversion to methylmercury (MeHg), a potent neurotoxin that bioaccumulates in food webs and poses risks to ecosystems and human health. Thiol functional groups (RSH) play an important role in controlling Hg(II) speciation and bio-uptake in methylating bacteria, yet the spatial distribution and density of these thiols within cells remain largely unknown. We isolated subcellular fractions of the Hg methylating bacterium Geobacter sulfurreducens in the exponential growth phase, and used Hg LIII-edge EXAFS (Extended X-ray Absorption Fine Structure) to quantify thiols in the extracellular medium, inner and outer membranes, periplasm and cytoplasm. The whole-cell thiol content was determined to be 1.3 × 10−10 μmol cell−1. The inner membrane contributed 7.1 × 10−11 (53%), the outer membrane 1.2 × 10−11 (9%), the periplasm 3.6 × 10−11 (27%) and the cytoplasm 1.5 × 10−11 μmol cell−1 (11%). The extracellular fraction contributed an additional 5.7 × 10−11 μmol cell−1, corresponding to 30% of the thiols of the cell culture. Local thiol density (thiols normalized to TOC in individual compartment, RSH/TOC, μmol g−1 C) was 36, 450, 140, 600 and 29 μmol g−1 C in the cytoplasm, inner membrane, periplasm, outer membrane and extracellular fractions, respectively. EXAFS analyses demonstrate Hg-thiolate coordination across all compartments, with Hg-O/N bonding and elemental Hg0 formed at higher Hg loadings. In the periplasm, Hg-disulfide and traces of β-HgS were detected. The high thiol density at the membranes, relative to other compartments, may imply they have an important role in the retention and internalization of Hg(II). Periplasmic thiols may modulate Hg(II) transfer between membranes, and cytoplasmic thiols may regulate the intracellular availability of Hg(II) for methylation. This work provides the first compartment-resolved quantification of thiol abundances and densities in a model Hg-methylating bacterium at subcellular level, offering a mechanistic framework for understanding the speciation, bioavailability, and subcellular transformation of Hg(II) with relevance for other soft metals (e.g., Cd, Pb, Zn, Ag, and Cu).
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Feb 2026
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I04-Macromolecular Crystallography
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Diamond Proposal Number(s):
[28402]
Open Access
Abstract: Despite the continual emergence of SARS-CoV-2 variants and increasing diversity within the receptor binding domain (RBD), some antibody responses that are directed to conserved regions can display cross-reactivity against variants. We previously isolated an RBD-directed monoclonal antibody (084-7D) from a Beta-infected donor that neutralized Beta and emerging Omicron variants. Here, we solved a high-resolution crystal structure of the 084-7D Fab in complex with the Beta RBD. These data revealed an epitope overlapping both the ACE2 binding site and those of other class 1 antibodies. Furthermore, the epitope includes highly conserved residues, Q409, D420, and Y489, that are present in recent Omicron variants. The N417 residue that emerged with Beta and has since persisted is tolerated within the epitope of 084-7D, explaining the preferential neutralization of contemporaneous N417-containing variants. These structural data defined the mechanism for cross-reactivity of a Beta-elicited neutralizing antibody, potentially informing the design of future broadly reactive SARS-CoV-2 therapeutics.
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Feb 2026
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