Native State Research Articles

Improved restoration of biomechanical factors using a narrow-box shaped reconstruction compared to a wide one in superior capsular reconstruction for irreparable supraspinatus tendon tears: a biomechanical study using a static shoulder simulator

PurposeTo biomechanically compare superior glenohumeral translation, subacromial contact pressures and area in a box-shape reconstruction using the long head of biceps tendon (LHBT) in an irreparable supraspinatus tendon tear model. MethodsSeven cadaveric shoulders (mean age 61 years; range 32-84 years; SD 22.3) were tested with a custom testing rig used to evaluate superior translation, subacromial contact pressures and areas at 0°, 30° and 60° of glenohumeral abduction. Conditions tested included the native state, a complete tear of the supraspinatus tendon, a wide box-shaped, double-bundle LHBT superior capsular reconstruction (wide BS-SCR), and a narrow box-shaped, double-bundle LHBT superior capsular reconstruction (narrow BS-SCR). ResultsCompared to the wide box-shape SCR, the narrow box-shape SCR had statistically significant lower median contact pressure at 30° and 60°. The subacromial contact area showed a statistically significant difference at 0° (p=0.001) and 30° (p=0.004) for the narrow compared to wide box-shape SCR. At an abduction angle of 0°, the narrow SCR could restore superior translation statistically significant better compared to the wide construct. For all angles, the wide and narrow box-shaped SCR increased the median subacromial distance statistically significantly. The contact areas in 30° and 60° of abduction were higher for all scenarios, both peaking in the intact state in 30° with approximately 600 mm2. ConclusionsIn comparison to a wide box-shape, a narrow box-shape SCR using the LHBT has biomechanical advantages in regard of subacromial contact pressures, the subacromial contact areas as well as the acromiohumeral distance. The width of the reconstruction therefore has direct influence in the success of the technique.

Transamniotic Stem Cell Therapy Modulates Uterine Natural Killer Cell Activity in the Hypoxia Model of Intrauterine Growth Restriction.

Intrauterine growth restriction (IUGR) pathophysiology is driven by abnormal uterine natural killer cell (uNK) activity leading to placental dysfunction. Transamniotic stem cell therapy (TRASCET) with mesenchymal stem cells (MSCs) can improve experimental IUGR by mechanisms not fully understood. We sought to examine TRASCET's effects in downstream products of uNKs in a model of IUGR: 15 Sprague-Dawley dams were exposed to alternating hypoxia (10.5% O2) from gestational day 15 (E15) until term (E21). Their fetuses (n = 189) were divided into four groups. One group remained untreated (n = 52), whereas three groups received volume-matched intraamniotic injections of either saline (sham, n = 44) or a suspension of amniotic fluid-derived MSCs, either in their native state (TRASCET, n = 50) or "primed" to an enhanced antiinflammatory phenotype (TRASCET-Primed, n = 43). Normal fetuses served as controls (n = 33). At term, various analyses were performed, including ELISA for surrogates of placental inflammation and uNK activity. Statistical comparisons included Bonferroni-adjusted criterion. Overall survival from hypoxia was 74% (140/189). Placental efficiency was lower in untreated and sham but normalized in both TRASCET groups (P < 0.01-0.47). Interleukin-17, a stimulator of uNKs, was elevated from normal in all groups (P < 0.01 for all). Interferon-gamma, released from activated uNKs, was elevated in all groups except sham but lower than the untreated in both TRASCET groups (P ≤ 0.01-0.06). Tumor necrosis factor-alpha, also produced by uNKs, was elevated in untreated and sham (P < 0.01 for both), but normalized by TRASCET (P = 0.05) and even lowered from normal in TRASCET-Primed (P < 0.01). Vascular endothelial growth factor, also released by uNKs, was elevated in untreated and sham but lower than normal in both TRASCET groups (P < 0.01 for all). We conclude that TRASCET with MSCs modulates the activity of placental uNKs in experimental IUGR, with distinct effects on their downstream products. This mechanistic insight may inform the development of novel strategies for the management of this disease.

Unlocking the unfolded structure of ubiquitin: Combining time-resolved x-ray solution scattering and molecular dynamics to generate unfolded ensembles.

The unfolding dynamics of ubiquitin were studied using a combination of x-ray solution scattering (XSS) and molecular dynamics (MD) simulations. The kinetic analysis of the XSS ubiquitin signals showed that the protein unfolds through a two-state process, independent of the presence of destabilizing salts. In order to characterize the ensemble of unfolded states in atomic detail, the experimental XSS results were used as a constraint in the MD simulations through the incorporation of x-ray scattering derived potential to drive the folded ubiquitin structure toward sampling unfolded states consistent with the XSS signals. We detail how biased MD simulations provide insight into unfolded states that are otherwise difficult to resolve and underscore how experimental XSS data can be combined with MD to efficiently sample structures away from the native state. Our results indicate that ubiquitin samples unfolded in states with a high degree of loss in secondary structure yet without a collapse to a molten globule or fully solvated extended chain. Finally, we propose how using biased-MD can significantly decrease the computational time and resources required to sample experimentally relevant nonequilibrium states.

Cryogenic electron microscopy reveals morphologically distinct subtypes of extracellular vesicles among porcine ejaculate fractions

Seminal plasma (SP) is rich in extracellular vesicles (EVs), which are still poorly studied, especially in livestock species. To better understand their functional role in both spermatozoa and endometrial epithelial cells, proper characterization of EVs is an essential step. The objective was to phenotypically characterize porcine seminal EVs (sEVs) using cryogenic electron microscopy (cryo-EM), which allows visualization of EVs in their native state. Porcine ejaculates are released in fractions, each containing SP from different source. This allows characterization sEVs released from various male reproductive tissues. Two experiments were performed, the first with SP from the entire ejaculate (n:6) and the second with SP from three ejaculate fractions (n:15): the first 10 mL of the sperm-rich ejaculate fraction (SRF-P1) with SP mainly from the epididymis, the remainder of the SRF (SRF-P2) with SP mainly from the prostate, and the post-SRF with SP mainly from the seminal vesicles. The sEVs were isolated by size exclusion chromatography and 1840 cryo-EM sEV images were acquired using a Jeol-JEM-2200FS/CR-EM. The size, electron density, complexity, and peripheral corona layer were measured in each sEV using the ImageJ software. The first experiment showed that sEVs were structurally and morphologically heterogeneous, although most (83.1%) were small (less than 200 nm), rounded, and poorly electrodense, and some have a peripheral coronal layer. There were also larger sEVs (16.9%) that were irregularly shaped, more electrodense, and few with a peripheral coronal layer. The second experiment showed that small sEVs were more common in SRF-P1 and SRF-P2, indicating that they originated mainly from the epididymis and prostate. Large sEVs were more abundant in post-SRF, indicating that they originated mainly from seminal vesicles. Porcine sEVs are structurally and morphologically heterogeneous. This would be explained by the diversity of reproductive organs of origin.

Reconstruction and identification of the native PLP synthase complex from Methanosarcina acetivorans lysate.

Many protein-protein interactions behave differently in biochemically purified forms as compared to their in vivo states. As such, determining native protein structures may elucidate structural states previously unknown for even well-characterized proteins. Here we apply the bottom-up structural proteomics method, cryoID , toward a model methanogenic archaeon. While they are keystone organisms in the global carbon cycle and active members of the human microbiome, there is a general lack of characterization of methanogen enzyme structure and function. Through the cryoID approach, we successfully reconstructed and identified the native Methanosarcina acetivorans pyridoxal 5'-phosphate (PLP) synthase (PdxS) complex directly from cryogenic electron microscopy (cryoEM) images of fractionated cellular lysate. We found that the native PdxS complex exists as a homo-dodecamer of PdxS subunits, and the previously proposed supracomplex containing both the synthase (PdxS) and glutaminase (PdxT) was not observed in cellular lysate. Our structure shows that the native PdxS monomer fashions a single 8α/8β TIM-barrel domain, surrounded by seven additional helices to mediate solvent and interface contacts. A density is present at the active site in the cryoEM map and is interpreted as ribose 5-phosphate. In addition to being the first reconstruction of the PdxS enzyme from a heterogeneous cellular sample, our results reveal a departure from previously published archaeal PdxS crystal structures, lacking the 37 amino acid insertion present in these prior cases. This study demonstrates the potential of applying the cryoID workflow to capture native structural states at atomic resolution for archaeal systems, for which traditional biochemical sample preparation is nontrivial.

Resolving chaperone-assisted protein folding on the ribosome at the peptide level.

Protein folding in vivo begins during synthesis on the ribosome and is modulated by molecular chaperones that engage the nascent polypeptide. How these features of protein biogenesis influence the maturation pathway of nascent proteins is incompletely understood. Here, we use hydrogen-deuterium exchange mass spectrometry to define, at peptide resolution, the cotranslational chaperone-assisted folding pathway of Escherichia coli dihydrofolate reductase. The nascent polypeptide folds along an unanticipated pathway through structured intermediates not populated during refolding from denaturant. Association with the ribosome allows these intermediates to form, as otherwise destabilizing carboxy-terminal sequences remain confined in the ribosome exit tunnel. Trigger factor binds partially folded states without disrupting their structure, and the nascent chain is poised to complete folding immediately upon emergence of the C terminus from the exit tunnel. By mapping interactions between the nascent chain and ribosomal proteins, we trace the path of the emerging polypeptide during synthesis. Our work reveals new mechanisms by which cellular factors shape the conformational search for the native state.

Amino-Acid Characteristics in Protein Native State Structures.

The molecular machines of life, proteins, are made up of twenty kinds of amino acids, each with distinctive side chains. We present a geometrical analysis of the protrusion statistics of side chains in more than 4000 high-resolution protein structures. We employ a coarse-grained representation of the protein backbone viewed as a linear chain of Cα atoms and consider just the heavy atoms of the side chains. We study the large variety of behaviors of the amino acids based on both rudimentary structural chemistry as well as geometry. Our geometrical analysis uses a backbone Frenet coordinate system for the common study of all amino acids. Our analysis underscores the richness of the repertoire of amino acids that is available to nature to design protein sequences that fit within the putative native state folds.

Structural details of helix-mediated TDP-43 C-terminal domain multimerization.

The primarily disordered C-terminal domain (CTD) of TAR DNA binding protein-43 (TDP-43), a key nuclear protein in RNA metabolism, forms neuronal inclusions in several neurodegenerative diseases. A conserved region (CR, spanning residues 319-341) in CTD forms transient helix-helix contacts important for its higher-order oligomerization and function that are disrupted by ALS-associated mutations. However, the structural details of CR assembly and the explanation for several ALS-associated variants' impact on phase separation and function remain unclear due to challenges in analyzing the dynamic association of TDP-43 CTD using traditional structural biology approaches. By employing an integrative approach, combining biophysical experiments, biochemical assays, AlphaFold2-Multimer (AF2-Multimer), and atomistic simulations, we generated structural models of helical oligomerization of TDP-43 CR. Using NMR, we first established that the native state of TDP-43 CR under physiological conditions is α-helical. Next, alanine scanning mutagenesis revealed that while hydrophobic residues in the CR are important for CR assembly, phase separation and TDP-43 nuclear retention function, polar residues down regulate these processes. Finally, pairing AF2-Multimer modeling with AAMD simulations indicated that dynamic, oligomeric assemblies of TDP-43 that are stabilized by a methionine-rich core with specific contributions from a tryptophan/leucine pair. In conclusion, our results advance the structural understanding of the mechanisms driving TDP-43 function and provide a window into the initial stages of its conversion into pathogenic aggregates.

The effect of heat treatment and high‐pressure homogenization on the dispersibility and interfacial behavior of faba bean protein isolate and concentrate

AbstractThe effect of hydrothermal treatments, high‐pressure homogenization (HPH) and their combined effects on faba bean protein isolate (FBI) and concentrate (FBC) dispersions was investigated. With HPH, a significant reduction in particle size was observed for FBI but not for FBC. For FBI, dispersion stability under accelerated gravitation was significantly improved by both hydrothermal and HPH. For FBC, hydrothermal treatment had a negative effect on dispersion stability unless it was combined with HPH. The protein dispersibility of FBI increased, and FBC decreased with increasing temperature. Upon HPH, the protein dispersibility of FBI dramatically improved, making it almost completely soluble. All FBI dispersions had a higher surface charge compared to FBC. SDS PAGE profile showed strong protein crosslinking in the original FBI, leading to a loss of legumin and vicilin bands, which were visible in a more native state of FBC. Physical modifications did not significantly affect the viscosities except for a significant increase in FBC heated at 75°C due to starch gelatinization. The equilibrium interfacial tension of the modified FBI was significantly lower than the FBC. Analysis of dynamic interfacial adsorption showed that FBI had a much faster initial diffusion (beyond detection limit) towards the interface than FBC. Physical modifications improved the surface hydrophobicity of the proteins, leading to faster interfacial adsorption. Overall, FBI dispersions had improved functional properties compared to FBC, and HPH had a more significant effect than hydrothermal treatments. Physically modified FBI could be a promising emulsifier in various food applications.

P-095 Towards deciphering the molecular architecture of the human sperm connecting piece

Abstract Study question How can we make the human sperm connecting piece accessible for structural analysis by cryo-electron tomography (cryo-ET)? Summary answer Preparation of sperm by chemical thinning or production of lamellae via cryo-focused ion beam (cryo-FIB) milling allows direct imaging of the human sperm connecting piece. What is known already Previous studies from our group and others have demonstrated the versatility of cryo-ET to advance our understanding of sperm biology. Structures of macromolecular complexes have been resolved in vitro and in situ, enabling a functional assessment at molecular or even atomic level. Resolving structures at high resolution further allowed identification of proteins previously not known to have a role in mediating male fertility. Such insights are instrumental to further improve diagnosis and treatment of male infertility. Study design, size, duration In our research study we use sperm samples from patients undergoing IVF treatment or donor material, which we receive through an ongoing collaboration with the Division of Woman and Baby, University Medical Centre Utrecht (UMCU). These samples are processed in our own laboratory and subsequently imaged by cryo-ET at the Electron Microscopy Centre of Utrecht University. Participants/materials, setting, methods Due to its thickness, the connecting piece cannot be directly analyzed by cryo-ET. Thus, we designed two strategies to achieve thinning of this area and allow collection of tomograms for structural analysis. While the first approach involves incubation of sperm with a detergent, a reducing agent and heparin, the second approach utilizes cryo-FIB milling to produce thin lamellae of the connecting piece. Main results and the role of chance Both strategies developed to access the connecting piece with cryo-ET are effective and allowed us a glimpse of the molecular landscape of the link between sperm head and tail. Chemical thinning of the connecting piece enabled imaging of the proximal and distal centriole while still embedded in the segmented columns or outer dense fibers, respectively. Furthermore, electron density within the microtubules of both centrioles indicates that they are decorated by microtubule inner proteins. Data obtained from lamellae produced by cryo-FIB milling complements these findings, displaying regular densities along the microtubules extending from the proximal centriole. Analyzing tomograms collected from cryo-FIB lamellae we further found a regular protein density that appears to be connecting the nuclear envelope to the base plate. Moreover, cryo-ET of lamellae allowed a detailed view of nuclear pore complexes and the postacrosomal sheath in their native environment. Limitations, reasons for caution Altering the native state of the sperm cell by chemical preparation is a limitation of this approach. Cryo-FIB milling retains the native state of the cell but is limited in throughput. To improve our cryo-FIB milling approach, we will implement automated milling procedures. Wider implications of the findings Chemical preparation and cryo-FIB milling of sperm enable the in-depth analysis of the connecting piece by cryo-ET. These strategies can now be applied to provide structural details at the molecular level and thus advance our understanding of male infertility related to defects of the head-tail junction. Trial registration number not applicable

Between Sādhū and Darwesh: Storytelling and Social Reform in S. uġhrā Humāyūñ Mirzā’s Mohinī (1929)

Ṣuġhrā Humāyūñ Mirzā (1884–1958) was a reformer, journalist, writer and traveller from the erstwhile princely state of Haidarābād, Dakkan. She wrote prolifically and used her fiction and non-fiction as a vehicle for her reformist ideas and objectives. However, scholarly focus on Ṣuġhrā Begum’s reformist work alone risks dismissal of her literary creativity and innovation. This paper aims to address this gap through an analysis of the narrative and stylistic features of Ṣuġhrā Begum’s Urdū novel, Mohinī (1929). I suggest a reading of this text on its own terms, i.e. one that emerges from the text and the motivations of its author, rather than through measurements with standard—usually Western—yardsticks or other assumptions about Ṣuġhrā Begum’s ostensibly singular priorities. Instead, I argue that Mohinī should be read in the context of the development of the Urdū novel, which was taking shape through the encounter between Western modes of representation made prominent through colonialism and existing Indian storytelling traditions, such as the qiṣṣāh. In the process, this paper expands the scholarship on this writer by offering a study of the creative features of her fiction and calls for the imagination of South Asian and wider Persianate print realisms that are not bound by rationalist epistemologies and encapsulate broader systems of knowledge.

The Yale-Cambridge Research Project to India: The Context, Disciplinary Character and Legal-Institutional Matters

This paper deals with the Yale-Cambridge Research Project which Helmut de Terra led to India in 1935. Its principal sponsors included the Carnegie Institution of Washington, Yale University and the American Philosophical Society. The expedition worked in various parts of British India and the princely state of Kashmir. The project is considered of exceptional importance in the field of both geological and archaeological research. This paper sets the historical context of the project and analyzes its disciplinary and scholarly character. The expedition aimed to address both geological and archaeological questions. In view of this interdisciplinary formation of the expedition, this study finally delves into legal and institutional matters so as to find out who were responsible for formally coordinating, such as issuance of the license etc. with de Terra and his team. The Geological Survey of India, the Archaeological Survey and the princely state of Kashmir, thus, come into our discussion.

Poster 187: Describing the Contribution of the Vacuum Effect and Hip Capsule in Restraint to Axial Traction in Patients Undergoing Hip Arthroscopy

Objectives: In recent investigations, there has been increased focus on hip laxity and hip capsular instability, particularly in the setting of hip arthroscopy. The capsule and its internal negative pressure effects have been shown to be major contributors to hip joint stability, however, the relative contributory role of these elements remains unknown. Intraoperatively, improved understanding of hip capsule biomechanics is imperative for optimization of arthroscopic technique to access the central compartment with minimal soft tissue trauma. In turn, this can help reduce the risk of hip instability and maximize patient outcomes. The purpose of this study was to define the stiffness coefficient of the hip capsule and vacuum phenomenon in response to hip axial traction. Our hypothesis was that the vacuum phenomenon would have a greater relative contribution to axial distraction stiffness at mid-range distraction, and that the hip capsule stiffness coefficient would be greater at terminal distraction. Methods: A review of a prospectively collected database of patients undergoing hip arthroscopy was conducted from 1 July 2021 to 31 June 2022. Patients in this database underwent hip arthroscopy for the treatment of femoroacetabular impingement syndrome (FAIS). Inclusion criteria were 1) completion of the study traction protocol and 2) completion of hip joint venting prior to capsulotomy. Exclusion criteria were 1) previous hip surgery and 2) lack of hip distraction at 50 pounds of axial traction. The study traction protocol involved applying axial traction prior to instrumentation (Native State) of the hip using a post-free distraction system. Hips were then vented using a spinal needle and the traction protocol was repeated (Vented State). Traction intervals were 0, 50, and 100 lbs of force. Fluoroscopic images were taken at each traction interval and standardized to preoperative radiographs, which allowed for a calculation of the lateral joint space width at each interval. Overall distraction distance was calculated as the difference between 0 lbs traction and 50 and 100 lbs traction. The stiffness coefficient was calculated as k = traction force interval/distraction distance in both the Native and Vented States. Comparisons using paired-samples t-tests were conducted between the stiffness coefficients of the Native State and Vented State at 50 lbs and at 100 lbs. A p-value of &lt;0.05 was utilized to indicate statistical significance. Results: Twenty-one patients were identified who met inclusion criteria. Mean age was 34.4 ± 13.4 years with 100% female and a mean BMI of 24.2 ± 4.8 points. Mean stiffness coefficient at 50 lbs in the Native State (10.9 ± 3.3) was significantly higher than the Vented State (4.9 ± 0.7, p &lt; 0.001), with similar findings at 100 lbs in the Native State (10.1 ± 1.7) and Vented State (8.7 ± 1.3, p &lt; 0.001). In the Vented State, the stiffness coefficient was significantly greater at 100 lbs (8.7 ± 1.3) than at 50 lbs (4.9 ± 0.7, p &lt; 0.001). Conclusions: Our results provide an objective characterization of the individual contributions of the hip soft tissue restraints and the vacuum phenomenon to hip resistance to axial traction. At 50 pounds of traction, the vacuum effect contributes to ~55% of the axial distraction stiffness (resistance to axial distraction). At 100 pounds of traction force, the vacuum effect contributes only ~14% of the axial distraction stiffness. The resistance of the hip to axial distraction is of substantial clinical significance as minimization of traction force decreases intraoperative soft tissue trauma. This, however, needs to be balanced with obtaining sufficient working space within the hip joint. There is still substantial controversy regarding the biomechanics of the hip and the optimal methods for minimizing traction force related injury (e.g., post-less traction, minimization of traction time, air arthrogram, and joint venting). These data demonstrate that soft tissues have a proportionally greater effect on restraint of the of the hip at terminal distraction (~1 cm distraction), whereas the vacuum effect has a greater influence on mid-range distraction (~0.5 cm distraction). The proposed model for this phenomenon is that the negative pressure and the soft tissue tension work in parallel to restrain the hip joint. The increased influence of the soft tissue tension at end-range distraction may be due to a check rein effect of the soft tissues; once the iliofemoral ligament has been tensioned to its maximum length its stiffness and capacity for transfer of tensile force increases rapidly. In conclusion, at 50 pounds of traction, the vacuum effect contributes to ~55% of the axial distraction stiffness (resistance to axial distraction). At 100 pounds of traction force, the vacuum effect contributes only ~14% of the axial distraction stiffness. These results demonstrate that the vacuum effect is a significant restraint to axial traction applied across the hip at mid-range distraction. Additionally, the hip capsule has a relatively greater contribution to terminal distraction stiffness.

Taylor-Aris Dispersion-Assisted Mass Spectrometry for the Analysis of Native Proteins.

As has recently been shown, Taylor-Aris dispersion-assisted mass spectrometry (TADA-MS) can offer direct injection MS determinations in fields where the targets of the analyses are large molecules present in a matrix that would otherwise cause serious interferences. In the present study, we demonstrated the exceptional utility of TADA-MS in native protein analysis: (i) a dramatic improvement in detection sensitivity was found due to its ability to strongly reduce matrix interferences, (ii) more "native-like" conditions can be used during analyses, (iii) the direct injection of non-MS-compatible matrices is allowed into MS, and (iv) a considerable simplification and economization of the workflow is ensured. We investigated the behavior of different types of proteins and protein complexes present under native conditions, demonstrating the unambiguous benefits and simplicity of the method.

Misleading Pore Size Measurements in Gelatin and Alginate Hydrogels Revealed by Confocal Microscopy.

It is a well-documented phenomenon that the porous structure of hydrogels observed with vacuum-based imaging techniques is generated during the freezing and drying process employed prior to observation. Nevertheless, vacuum-based techniques, such as scanning electron microscopy (SEM), are still being commonly used to measure pore sizes in hydrogels, which is often not representative of the actual pore size in hydrated conditions. The frequent underestimation of the impact of freezing and drying on hydrogel structures could stem from a lack of cross-fertilization between materials science and biomedical or food science communities, or from the simplicity and visually appealing nature of SEM imaging, which may lead to an overemphasis on its use. Our study provides a straightforward and impactful way of pinpointing this phenomenon exploiting two hydrogels ubiquitously applied in tissue engineering, including gelatin methacryloyl and alginate as proof-of-concept hydrogels. By comparing images of the samples in the native hydrated state, followed by freezing, freeze-drying, and rehydration using SEM and confocal microscopy, we highlight discrepancies between hydrogel pore sizes in the hydrated versus the dry state. To conclude, our study offers recommendations for researchers seeking insight in hydrogel properties and emphasizes key factors that require careful control when using SEM as a characterization tool.

Biomechanical evaluation of spinal cord decompression methods in dogs

The study is relevant due to the high incidence of spinal cord compression pathologies in dogs requiring surgical intervention, and was carried out with the aim of conducting a biomechanical assessment of spinal cord decompression methods and identifying a decompression technique that has the least destabilizing effect on the thoracolumbar spine. The studies were carried out in 2023-2024. on the basis of the Department of Veterinary Surgery of the Moscow State Academy of Veterinary Medicine and Biotechnology - MBA named after K.I. Scriabin" and the Laboratory for the Development and Testing of Medical Devices and Materials of the Federal State Budgetary Institution "National Medical Research Center for Orthopedics named after. N.N. Priorov" of the Ministry of Health of the Russian Federation. The object of the study was sectional material, namely, samples of the thoracolumbar spine of dogs of chondrodystrophic and non-chondrodystrophic breeds weighing from 3 to 10 kg at the age of 5 to 8 years (20 samples in total). The range of motion of the vertebrae and the force of destruction of the spinal motion segments were determined in the native state and after modeling decompression techniques (hemilaminectomy and mini-hemilaminectomy). During hemilaminectomy, statistically significant changes in the biomechanical characteristics of the thoracolumbar spine were noted compared to mini-hemilaminectomy: an increase in the range of motion of the vertebrae in the area of decompression; increasing the range of motion of the vertebrae in topographically adjacent segments; reduction in the force of destruction of the segment in the area of decompression (changes are more pronounced in dogs of chondrodystrophic breeds). The data obtained by the authors of the article indicate that hemilaminectomy is a more destabilizing technique compared to minihemilaminectomy, and for extrusions in the thoracolumbar spine in dogs, preference should be given to mini-hemilaminectomy.

A systematic evaluation of physiochemical properties and solar-driven photocatalytic activity of nanosized Mn doped CdS on Methylene Blue dye

Manganese ions were incorporated into CdS nanosystem using wet chemical protocol at different pH values namely 9.0,10.0 and 11.0 and the same were used for photocatalytic degradation of Methylene blue (MB) dye via adsorption phenomena. Several standard analytical techniques were employed to analyse the synthesized nanocompounds: XRD, scanning electron microscopy with energy-dispersive X-ray spectroscopy, transmission electron microscopy, UV-Vis absorption spectroscopy, Fourier transform infrared spectroscopy, and vibrating-sample magnetometry. The structural analysis revealed the presence of primary cubic zinc blende structure with secondary phases. The morphological studies confirmed the formation of nanostructured particles for all pH values, amongst, pH 10.0 grown particles were showing less agglomeration and better particle stabilization. The TEM analysis confirmed the particle size of 23 nm. The absorption edge and its position shown by UV-Vis absorption analysis exhibited a broad envelope and strong blue shift due to an uneven size distribution and nanodimensional state formation of particles, respectively. The elevation to superparamagnetic state from native diamagnetic state of CdS due to Mn ions incorporation was confirmed by VSM studies. The dual phase composition significantly enhanced the photocatalytic efficiency by improving charge separation and extending light absorption. The MnS phase effectively trapped electrons, reduced recombination rates and facilitated the generation of reactive species. The photocatalytic degradation of methylene blue dye was more efficient (88.87 %) under direct sunlight exposure. Additionally, the enhanced surface properties and adsorption characteristics, as confirmed by the Freundlich isotherm and intraparticle diffusion model, contributed to the superior degradation performance.

Cysteine Oxidation in Human Galectin-1 Occurs Sequentially via a Folded Intermediate to a Fully Oxidized Unfolded Form.

Galectins are multifunctional effectors in cellular homeostasis and dysregulation. Oxidation of human galectin-1 (Gal-1) with its six sulfhydryls produces a disulfide-bridged oxidized form that lacks normal lectin activity yet gains new glycan-independent functionality. Nevertheless, the mechanistic details as to how Gal-1 oxidation occurs remain unclear. Here, we used 15N and 13C HSQC NMR spectroscopy to gain structural insight into the CuSO4-mediated path of Gal-1 oxidation and identified a minimum two-stage conversion process. During the first phase, disulfide bridges form slowly between C16-C88 and/or C42-C66 to produce a partially oxidized, conformationally flexible intermediate that retains the ability to bind lactose. Site-directed mutagenesis of C16 to S16 impedes the onset of this overall slow process. During the second phase, increased motional dynamics of the intermediate enable the relatively distant C2 and C130 residues to form the third and final disulfide bond, leading to an unfolded state and consequent dimer dissociation. This fully oxidized end state loses the ability to bind lactose, as shown by the hemagglutination assay. Consistent with this model, we observed that the Gal-1 C2S mutant maintains intermediate-state structural features with a free sulfhydryl group at C130. Incubation with dithiothreitol reduces all disulfide bonds and allows the lectin to revert to its native state. Thus, the sequential, non-random formation of three disulfide bridges in Gal-1 in an oxidative environment acts as a molecular switch for fundamental changes to its functionality. These data inspire detailed bioactivity analysis of the structurally defined oxidized intermediate in, e.g., acute and chronic inflammation.

The Structural Stability of Membrane Proteins Revisited: Combined Thermodynamic and Spectral Phasor Analysis of SDS-induced Denaturation of a Thermophilic Cu(I)-transport ATPase

Assessing membrane protein stability is among the major challenges in protein science due to their inherent complexity, which complicates the application of conventional biophysical tools. In this work, sodium dodecyl sulfate-induced denaturation of AfCopA, a Cu(I)-transport ATPase from Archaeoglobus fulgidus, was explored using a combined model-free spectral phasor analysis and a model-dependent thermodynamic analysis. Decrease in tryptophan and 1-anilino-naphthalene-8-sulfonate fluorescence intensity, displacements in the spectral phasor space, and the loss of ATPase activity were reversibly induced by this detergent. Refolding from the SDS-induced denatured state yields an active enzyme that is functionally and spectroscopically indistinguishable from the native state of the protein. Phasor analysis of Trp spectra allowed us to identify two intermediate states in the SDS-induced denaturation of AfCopA, a result further supported by principal component analysis. In contrast, traditional thermodynamic analysis detected only one intermediate state, and including the second one led to overparameterization. Additionally, ANS fluorescence spectral analysis detected one more intermediate and a gradual change at the level of the hydrophobic transmembrane surface of the protein. Based on this evidence, a model for acquiring the native structure of AfCopA in a membrane-like environment is proposed.

A life in light - in honor of David Mauzerall on his 95th birthday.

David Mauzerall was born on July 22, 1929 to a working-class family in the small, inland textile town of Sanford, Maine. Those humble origins instilled a lifelong frugality and an innovative spirit. After earning his PhD degree in 1954 in physical organic chemistry with Frank Westheimer at the University of Chicago, he joined The Rockefeller Institute for Medical Research (now University) as a postdoctoral fellow that summer, rose to the rank of professor, and remained there for the rest of his career. His work over more than 60 years encompassed porphyrin biosynthesis, photoinduced electron-transfer reactions in diverse architectures (solutions, bilayer lipid membranes, reaction centers, chromatophores, and intact leaves), the light-saturation curve of photosynthesis, statistical treatments of photoreactions, and "all-things porphyrins." His research culminated in studies he poetically referred to as "listening to leaves" through the use of pulsed photoacoustic spectroscopy to probe the course and thermodynamics of photosynthesis in its native state. His research group was always small; indeed, of 185 total publications, 39 were singly authored. In brief, David Mauzerall has blended a deep knowledge of distinct disciplines of physical organic chemistry, photochemistry, spectroscopy and biophysics with ingenious experimental methods, incisive mathematical analysis, pristine personal integrity, and unyielding love of science to deepen our understanding of photosynthesis in its broadest context. He thought creatively - and always independently. His work helped systematize the fields of photosynthesis and the origin of life and made them more quantitative. The present article highlights a number of salient scientific discoveries and includes comments from members of his family, friends, and collaborators (Gary Brudvig, Greg Edens, Paul Falkowski, Alzatta Fogg, G. Govindjee, Nancy Greenbaum, Marilyn Gunner, Harvey Hou, Denise and Michele Mauzerall, Thomas Moore, and William Parson) as part of a celebration of his 95th birthday.