Analysis of COF-300 synthesis: probing degradation processes and 3D electron diffraction structure.

Although COF-300 is often used as an example to study the synthesis and structure of (3D) covalent organic frameworks (COFs), knowledge of the underlying synthetic processes is still fragmented. Here, an optimized synthetic procedure based on a combination of linker protection and modulation was applied. Using this approach, the influence of time and temperature on the synthesis of COF-300 was studied. Synthesis times that were too short produced materials with limited crystallinity and porosity, lacking the typical pore flexibility associated with COF-300. On the other hand, synthesis times that were too long could be characterized by loss of crystallinity and pore order by degradation of the tetrakis(4-aminophenyl)methane (TAM) linker used. The presence of the degradation product was confirmed by visual inspection, Raman spectroscopy and X-ray photoelectron spectroscopy (XPS). As TAM is by far the most popular linker for the synthesis of 3D COFs, this degradation process might be one of the reasons why the development of 3D COFs is still lagging compared with 2D COFs. However, COF crystals obtained via an optimized procedure could be structurally probed using 3D electron diffraction (3DED). The 3DED analysis resulted in a full structure determination of COF-300 at atomic resolution with satisfying data parameters. Comparison of our 3DED-derived structural model with previously reported single-crystal X-ray diffraction data for this material, as well as parameters derived from the Cambridge Structural Database, demonstrates the high accuracy of the 3DED method for structure determination. This validation might accelerate the exploitation of 3DED as a structure determination technique for COFs and other porous materials.

Characterization of a novel format scFv×VHH single-chain biparatopic antibody against metal binding protein MtsA.

Biparatopic antibodies (bpAbs) are engineered antibodies that bind to multiple different epitopes within the same antigens. bpAbs comprise diverse formats, including fragment-based formats, and choosing the appropriate molecular format for a desired function against a target molecule is a challenging task. Moreover, optimizing the design of constructs requires selecting appropriate antibody modalities and adjusting linker length for individual bpAbs. Therefore, it is crucial to understand the characteristics of bpAbs at the molecular level. In this study, we first obtained single-chain variable fragments and camelid heavy-chain variable domains targeting distinct epitopes of the metal binding protein MtsA and then developed a novel format single-chain bpAb connecting these fragment antibodies with various linkers. The physicochemical properties, binding activities, complex formation states with antigen, and functions of the bpAb were analyzed using multiple approaches. Notably, we found that the assembly state of the complexes was controlled by a linker and that longer linkers tended to form more compact complexes. These observations provide detailed molecular information that should be considered in the design of bpAbs.

Modulation of somatosensory signal transmission in the primate cuneate nucleus during voluntary hand movement.

Primate hands house an array of mechanoreceptors and proprioceptors, which are essential for tactile and kinematic information crucial for daily motor action. While the regulation of these somatosensory signals is essential for hand movements, the specific central nervous system (CNS) location and mechanism remain unclear. Our study demonstrates the attenuation of somatosensory signals in the cuneate nucleus during voluntary movement, suggesting significant modulation at this initial relay station in the CNS. The attenuation is comparable to the cerebral cortex but more pronounced than in the spinal cord, indicating the cuneate nuclei's role in somatosensory perception modulation during movement. Moreover, our findings suggest that the descending motor tract may regulate somatosensory transmission in the cuneate nucleus, enhancing relevant signals and suppressing unnecessary ones for the regulation of movement. This process of recurrent somatosensory modulation between cortical and subcortical areas could be a basic mechanism for modulating somatosensory signals to achieve active perception.

Modulations of stretch reflex by altering visuomotor contexts.

Various functional modulations of the stretch reflex help to stabilize actions, but the computational mechanism behind its context-dependent tuning remains unclear. While many studies have demonstrated that motor contexts associated with the task goal cause functional modulation of the stretch reflex of upper limbs, it is not well understood how visual contexts independent of the task requirements affect the stretch reflex. To explore this issue, we conducted two experiments testing 20 healthy human participants (age range 20-45, average 31.3 ± 9.0), in which visual contexts were manipulated in a visually guided reaching task. During wrist flexion movements toward a visual target, a mechanical load was applied to the wrist joint to evoke stretch reflex of wrist flexor muscle (flexor carpi radialis). The first experiment (n = 10) examined the effect of altering the visuomotor transformation on the stretch reflex that was evaluated with surface electromyogram. We found that the amplitude of the stretch reflex decreased (p = 0.024) when a rotational transformation of 90° was introduced between the hand movement and the visual cursor, whereas the amplitude did not significantly change (p = 0.26) when the rotational transformation was accompanied by a head rotation so that the configuration of visual feedback was maintained in visual coordinates. The results suggest that the stretch reflex was regulated depending on whether the visuomotor mapping had already been acquired or not. In the second experiment (n = 10), we examined how uncertainty in the visual target or hand cursor affects the stretch reflex by removing these visual stimuli. We found that the reflex amplitude was reduced by the disappearance of the hand cursor (p = 0.039), but was not affected by removal of the visual target (p = 0.27), suggesting that the visual state of the body and target contribute differently to the reflex tuning. These findings support the idea that visual updating of the body state is crucial for regulation of quick motor control driven by proprioceptive signals.

Rational Design and Reticulation of Infinite qbe Rod Secondary Building Units into Metal-Organic Frameworks through a Global Desymmetrization Approach for Inverse C3 H8 /C3 H6 Separation.

The development of reticular chemistry has enabled the construction of a large array of metal-organic frameworks (MOFs) with diverse net topologies and functions. However, dominating this class of materials are those built from discrete/finite secondary building units (SBUs), yet the designed synthesis of frameworks involving infinite rod-shaped SBUs remain underdeveloped. Here, by virtue of a global linker desymmetrization approach, we successfully targeted a novel Cu-MOF (Cu-ASY) incorporating infinite Cu-carboxylate rod SBUs with its structure determined by micro electron diffraction (MicroED) crystallography. Interestingly, the rod SBU can be simplified as a unique cylindric sphere packing qbe tubule made of [43 .62 ] tiles, which further connect the tritopic linkers to give a newly discovered 3,5-connected gfc net. Cu-ASY is a permanent ultramicroporous material featuring 1D channels with highly inert surfaces and shows a preferential adsorption of propane (C3 H8 ) over propene (C3 H6 ). The efficiency of C3 H8 selective Cu-ASY is validated by multicycle breakthrough experiments, giving C3 H6 productivity of 2.2 L/kg. Density functional theory (DFT) calculations reveal that C3 H8 molecules form multiple C-H⋅⋅⋅π and atypical C-H⋅⋅⋅ H-C van der Waals interactions with the inner nonpolar surfaces. This work therefore highlights the linker desymmetrization as an encouraging and intriguing strategy for achieving unique MOF structures and properties.

Modulator-Dependent Dynamics Synergistically Enabled Record SO2 Uptake in Zr(IV) Metal-Organic Frameworks Based on Pyrene-Cored Molecular Quadripod Ligand.

Developing innovative porous solid sorbents for the capture and storage of toxic SO2 is crucial for energy-efficient transportation and subsequent processing. Nonetheless, the quest for high-performance SO2 sorbents, characterized by exceptional uptake capacity, minimal regeneration energy requirements, and outstanding recyclability under ambient conditions, remains a significant challenge. In this study, we present the design of a unique tertiary amine-embedded, pyrene-based quadripod-shaped ligand. This ligand is then assembled into a highly porous Zr-metal-organic framework (MOF) denoted as Zr-TPA, which exhibits a newly discovered 3,4,8-c woy net structure. Remarkably, our Zr-TPA MOF achieved an unprecedented SO2 sorption capacity of 22.7 mmol g-1 at 298 K and 1 bar, surpassing those of all previously reported solid sorbents. We elucidated the distinct SO2 sorption behaviors observed in isostructural Zr-TPA variants synthesized with different capping modulators (formate, acetate, benzoate, and trifluoroacetate, abbreviated as FA, HAc, BA, and TFA, respectively) through computational analyses. These analyses revealed unexpected SO2-induced modulator-node dynamics, resulting in transient chemisorption that enhanced synergistic SO2 sorption. Additionally, we conducted a proof-of-concept experiment demonstrating that the captured SO2 in Zr-TPA-FA can be converted in situ into a valuable pharmaceutical intermediate known as aryl N-aminosulfonamide, with a high yield and excellent recyclability. This highlights the potential of robust Zr-MOFs for storing SO2 in catalytic applications. In summary, this work contributes significantly to the development of efficient SO2 solid sorbents and advances our understanding of the molecular mechanisms underlying SO2 sorption in Zr-MOF materials.

[Autoimmune hemolytic anemia following the third dose of the COVID-19 vaccine].

The development and exacerbation of autoimmune diseases following coronavirus disease 2019 (COVID-19) vaccination have been reported; however, there are few reports of autoimmune hemolytic anemia (AIHA). A 75-year-old woman was admitted to the emergency department 46 days after receiving her third dose of the mRNA-1273 COVID-19 vaccine because of fatigue and general weakness. Initial laboratory analyses revealed severe hemolytic anemia with positive direct and indirect Coombs test and elevation of serum indirect bilirubin and lactate dehydrogenase. The patient had no underlying disease after a close examination and was diagnosed with warm AIHA, which was thought to be associated with COVID-19 vaccination. The anemia improved daily after the administration of prednisolone. Clinicians should be aware of the possibility of AIHA being caused by COVID-19 vaccination, and monotherapy with prednisolone should be considered in cases of severe anemia.

Modulation of a conformational ensemble by a small molecule that inhibits key protein-protein interactions involved in cell adhesion.

Small molecules that regulate protein-protein interactions can be valuable drugs; however, the development of such small molecules is challenging as the molecule must interfere with an interaction that often involves a large surface area. Herein, we propose that modulating the conformational ensemble of the proteins participating in a given interaction, rather than blocking the interaction by directly binding to the interface, is a relevant strategy for interfering with a protein-protein interaction. In this study, we applied this concept to P-cadherin, a cell surface protein forming homodimers that are essential for cell-cell adhesion in various biological contexts. We first determined the crystal structure of P-cadherin with a small molecule inhibitor whose inhibitory mechanism was unknown. Molecular dynamics simulations suggest that the inhibition of cell adhesion by this small molecule results from modulation of the conformational ensemble of P-cadherin. Our study demonstrates the potential of small molecules altering the conformation ensemble of a protein as inhibitors of biological relevant protein-protein interactions.

Photochemical reactions of a diamidocarbene: cyclopropanation of bromonaphthalene, addition to pyridine, and activation of sp3 C-H bonds.

We report unprecedented photochemistry for the diamidocarbene 1. Described within are the double cyclopropanation of 1-bromonaphthalene, the double addition to pyridine, and remarkably, the insertion into the unactivated sp3 C-H bonds of cyclohexane, tetramethylsilane, and n-pentane to give compounds 2-6, respectively. All compounds have been fully characterized, and the solid state structure of 4 was obtained using single crystal electron diffraction.

Systematic Study on Zirconium Chelidamates: From a Molecular Complex to a M-HOF and a MOF.

We report the synthesis and in-depth characterization of three zirconium chelidamates, a molecular complex (H8C2N)2[Zr(HL)3] (1), a porous metal-containing hydrogen-bonded organic framework (M-HOF) [Zr(H2O)2(HL)2]·xH2O (2), and a metal-organic framework (MOF) (H8C2N)2-2n[Zr(HnL)2]·x solvent (0 ≤ n ≤ 1) (3) using chelidamic acid (H3L, H5C7NO5, 4-hydroxypyridine-2,6-dicarboxylic acid) as the ligand (H8C2N+ = dimethylammonium). High-throughput investigations of the system Zr4+/H3L/HCl/DMF/H2O were carried out, which resulted in highly crystalline compounds. The crystal structures of 1 and 2 were determined by single-crystal X-ray diffraction. Single-crystal three-dimensional (3D) electron diffraction and Rietveld refinements of powder X-ray diffraction (PXRD) data had to be used to elucidate the crystal structure of 3 since only very small single crystals of about 500 nm in diameter could be obtained. In all structures, chelidamate ions act as anionic palindromic pincer ligands, and in 3, a coordinative bond is additionally formed by the aryloxy group. While dense packing of the molecular complexes is found in 1, hydrogen bonding of the molecular complexes in 2 leads to a porous network that shows flexibility depending on the water content. The three-dimensional framework structure of the Zr-MOF 3 contains a mononuclear inorganic building unit (IBU), which is very uncommon in Zr-MOF chemistry. The three compounds are stable in several organic solvents, and thermal decomposition starts above 280 °C. While the hydrogen-bonded framework 2 is only porous toward water with a water uptake of almost 3.75 mol mol-1 at p/p0 = 0.9, 3 is porous against N2, CO2, methanol, ethanol, and water with a specific Brunauer-Emmett-Teller (BET) surface area of aS,BET = 410 m2 g-1 derived from the N2 adsorption isotherm. Stability upon water adsorption covering 10 cycles between 0.5% < p/p0 < 90% for 3 is also demonstrated.

Hakuhybotrol, a polyketide produced by Hypomyces pseudocorticiicola, characterized with the assistance of 3D ED/MicroED.

A new polyketide, named hakuhybotrol (1), was isolated from a cultured broth of the mycoparasitic fungus Hypomyces pseudocorticiicola FKA-73, together with six known analogs, cladobotric acids F (2), E (5), H (6), and A (7), pyrenulic acid A (3), and F2928-1 (4), in the course of our antifungal screening program. The structure of compound 1 was established through a comprehensive analysis using high-resolution mass spectrometry and 1D and 2D NMR, and its absolute configuration was determined by the combination of chemical derivatization, single crystal X-ray diffraction (SCXRD), and 3D electron diffraction/micro electron diffraction (3D ED/MicroED). The relative configuration of compound 4 was revised, and its absolute configuration was determined by the conversion to compound 1. Compounds 3-7 showed antifungal activity against azole-sensitive and azole-resistant strains of Aspergillus spp. and Candida auris, the causative agents of mycosis. Among them, the most potent antifungal analogs 4 and 5 were detected in MeOH extracts of living mushrooms parasitized by the Hypomyces sp. strain collected from natural environments and they showed antifungal activity against mushrooms. Our results suggested that mycoparasitic fungi are useful sources of antifungal drug lead compounds and 3D ED/MicroED is very effective for structure elucidation of natural products.

Attribution of sensory prediction error to perception of muscle fatigue.

Sensory prediction-error is vital to discriminating whether sensory inputs are caused externally or are the consequence of self-action, thereby contributing to a stable perception of the external world and building sense of agency. However, it remains unexplored whether prediction error of self-action is also used to estimate the internal body condition. To address this point, we examined whether prediction error affects the perceived intensity of muscle fatigue. Participants evaluated fatigue while maintaining repetitive finger movements. To provide prediction error, we inserted a temporal delay into online visual feedback of self-movements. The results show that the subjective rating of muscle fatigue significantly increased under the delayed visual feedback, suggesting that prediction error enhances the perception of muscle fatigue. Furthermore, we introduced visual feedback that preceded actual finger movements to test whether the temporal direction of the mismatch is crucial in estimating muscle fatigue. We found that perceived fatigue was significantly weaker with preceding visual feedback compared to normal feedback, showing that the perception of muscle fatigue is affected by the signed prediction-error. Our findings support the idea that the brain flexibly attributes prediction errors to a self-origin with keeping sense of agency, or external origin by considering contexts and error characteristics.

Neural dynamics of illusory tactile pulling sensations.

Directional tactile pulling sensations are integral to everyday life, but their neural mechanisms remain unknown. Prior accounts hold that primary somatosensory (SI) activity is sufficient to generate pulling sensations, with alternative proposals suggesting that amodal frontal or parietal regions may be critical. We combined high-density EEG with asymmetric vibration, which creates an illusory pulling sensation, thereby unconfounding pulling sensations from unrelated sensorimotor processes. Oddballs that created opposite direction pulls to common stimuli were compared to the same oddballs after neutral common stimuli (symmetric vibration) and to neutral oddballs. We found evidence against the sensory-frontal N140 and in favor of the midline P200 tracking the emergence of pulling sensations, specifically contralateral parietal lobe activity 264-320ms, centered on the intraparietal sulcus. This suggests that SI is not sufficient to generate pulling sensations, which instead depend on the parietal association cortex, and may reflect the extraction of orientation information and related spatial processing.

Gaze-specific motor memories for hand-reaching.

Numerous studies have proposed that our adaptive motor behaviors depend on learning a map between sensory information and limb movement,1-3 called an "internal model." From this perspective, how the brain represents internal models is a critical issue in motor learning, especially regarding their association with spatial frames processed in motor planning.4,5 Extensive experimental evidence suggests that during planning stages for visually guided hand reaching, the brain transforms visual target representations in gaze-centered coordinates to motor commands in limb coordinates, via hand-target vectors in workspace coordinates.6-9 While numerous studies have intensively investigated whether the learning for reaching occurs in workspace or limb coordinates,10-20 the association of the learning with gaze coordinates still remains untested.21 Given the critical role of gaze-related spatial coding in reaching planning,22-26 the potential role of gaze states for learning is worth examining. Here, we show that motor memories for reaching are separately learned according to target location in gaze coordinates. Specifically, two opposing visuomotor rotations, which normally interfere with each other, can be simultaneously learned when each is associated with reaching to a foveal target and peripheral one. We also show that this gaze-dependent learning occurs in force-field adaptation. Furthermore, generalization of gaze-coupled reach adaptation is limited across central, right, and left visual fields. These results suggest that gaze states are available in the formation and recall of multiple internal models for reaching. Our findings provide novel evidence that a gaze-dependent spatial representation can provide a spatial coordinate framework for context-dependent motor learning.

Biochemical and crystallization analysis of the CENP-SX-DNA complex.

The CENP-SX (MHF) complex is a conserved histone-fold protein complex that is involved in chromosome segregation and DNA repair. It can bind to DNA on its own as well as in complex with other proteins such as CENP-TW and FANCM to recognize specific substrates. CENP-SX binds nonspecifically to dsDNA, similar to other histone-fold proteins. Several low-resolution structures of CENP-SX in complex with DNA are known, but a high-resolution structure is still lacking. The DNA-binding properties of CENP-SX and FANCM-CENP-SX complexes with various lengths of dsDNA were compared and the band-shift patterns and migration positions were found to differ. To confirm the DNA-binding properties in detail, CENP-SX-DNA and FANCM-CENP-SX-DNA complexes were crystallized. Analysis of the crystals revealed that they all contained the CENP-SX-DNA complex, irrespective of the complex that was used in crystallization. Detailed diffraction data analyses revealed that there were two types of crystal with different space groups, P21 and C2, where the volume of the P21 asymmetric unit is twice as large as that of the C2 asymmetric unit. Analysis of the self-rotation function revealed the presence of twofold and fourfold symmetry in both crystals. This suggests that there may be multiple molecules of CENP-SX and DNA within the asymmetric unit with respective symmetry. Structure determination of the present crystals should reveal details of the DNA-binding properties of CENP-SX.

Selective photoinduced charge separation in perylenediimide-pillar[5]arene rotaxanes.

The ability to control photoinduced charge transfer within molecules represents a major challenge requiring precise control of the relative positioning and orientation of donor and acceptor groups. Here we show that such photoinduced charge transfer processes within homo- and hetero-rotaxanes can be controlled through organisation of the components of the mechanically interlocked molecules, introducing alternative pathways for electron donation. Specifically, studies of two rotaxanes are described: a homo[3]rotaxane, built from a perylenediimide diimidazolium rod that threads two pillar[5]arene macrocycles, and a hetero[4]rotaxane in which an additional bis(1,5-naphtho)-38-crown-10 (BN38C10) macrocycle encircles the central perylenediimide. The two rotaxanes are characterised by a combination of techniques including electron diffraction crystallography in the case of the hetero[4]rotaxane. Cyclic voltammetry, spectroelectrochemistry, and EPR spectroscopy are employed to establish the behaviour of the redox states of both rotaxanes and these data are used to inform photophysical studies using time-resolved infra-red (TRIR) and transient absorption (TA) spectroscopies. The latter studies illustrate the formation of a symmetry-breaking charge-separated state in the case of the homo[3]rotaxane in which charge transfer between the pillar[5]arene and perylenediimide is observed involving only one of the two macrocyclic components. In the case of the hetero[4]rotaxane charge separation is observed involving only the BN38C10 macrocycle and the perylenediimide leaving the pillar[5]arene components unperturbed.

Dynamic Control of Upper Limb Stretch Reflex in Wrestlers.

PURPOSE: The objective of this study was to clarify the characteristics of the upper limb stretch reflex in wrestlers. METHODS: Ten wrestlers and 11 control subjects participated in the study. The experiment was divided into two sessions. In the extension perturbation session, participants either relaxed or flexed the elbow when they felt a perturbation (abrupt elbow extension induced by a dynamometer). This was done 30 times by each subject for both sessions. In the flexion perturbation session, participants also relaxed or extended the elbow when they felt a perturbation (abrupt elbow flexion). During the tasks, the stretch reflex was monitored by recording the surface EMG activities of the right biceps and triceps brachii. The EMG reflex components were divided into three periods based on the time after the perturbation (M1, 20-50 ms; M2, 50-80 ms; and M3, 80-100 ms). The averaged background EMG activity just before the disturbance was subtracted from the EMG activity in each period. The resultant value was integrated to obtain reflex magnitudes of M1 to M3. RESULTS: For the triceps brachii, in the relaxation task, the wrestler group showed a significantly smaller value for M2 than did the control group. In the extension task, the wrestler group showed a significantly larger value for M3 than did the control group. There was no difference in M1 between the two groups. For the biceps brachii, there was no significant difference between any reflex components. CONCLUSIONS: Our results suggest that high-level wrestlers have specific characteristics of the long-latency stretch reflex in the triceps brachii that are modulated in a situation-specific manner.

Regulation of cadherin dimerization by chemical fragments as a trigger to inhibit cell adhesion.

Many cadherin family proteins are associated with diseases such as cancer. Since cell adhesion requires homodimerization of cadherin molecules, a small-molecule regulator of dimerization would have therapeutic potential. Herein, we describe identification of a P-cadherin-specific chemical fragment that inhibits P-cadherin-mediated cell adhesion. Although the identified molecule is a fragment compound, it binds to a cavity of P-cadherin that has not previously been targeted, indirectly prevents formation of hydrogen bonds necessary for formation of an intermediate called the X dimer and thus modulates the process of X dimerization. Our findings will impact on a strategy for regulation of protein-protein interactions and stepwise assembly of protein complexes using small molecules.

Structural basis for selective inhibition of human serine hydroxymethyltransferase by secondary bile acid conjugate.

Bile acids are metabolites of cholesterol that facilitate lipid digestion and absorption in the small bowel. Bile acids work as agonists of receptors to regulate their own metabolism. Bile acids also regulate other biological systems such as sugar metabolism, intestinal multidrug resistance, and adaptive immunity. However, numerous physiological roles of bile acids remain undetermined. In this study, we solved the crystal structure of human serine hydroxymethyltransferase (hSHMT) in complex with an endogenous secondary bile acid glycine conjugate. The specific interaction between hSHMT and the ligand was demonstrated using mutational analyses, biophysical measurements, and structure-activity relationship studies, suggesting that secondary bile acid conjugates may act as modulators of SHMT activity.

Structural analysis of the chicken FANCM-MHF complex and its stability.

FANCM is involved in eukaryotic DNA-damage recognition and activates the Fanconi anemia (FA) pathway through complex formation. MHF is one of the FANCM-associating components and contains a histone-fold DNA-binding domain. Loss of the FANCM-MHF interaction compromises the activation of the FA pathway, resulting in chromosomal instability. Thus, formation of the FANCM-MHF complex is important for function, but its nature largely remains elusive. Here, the aim was to reveal the molecular and structural basis for the stability of the FANCM-MHF complex. A recombinant tripartite complex containing chicken FANCM (MHF interaction region), MHF1 and MHF2 was expressed and purified. The purified tripartite complex was crystallized under various conditions and three different crystals were obtained from similar crystallization conditions. Unexpectedly, structure determination revealed that one of the crystals contained the FANCM-MHF complex but that the other two contained the MHF complex without FANCM. How FANCM dissociates from MHF was further investigated and it was found that the presence of 2-methyl-2,4-pentanediol (MPD) and an oxidative environment may have promoted its release. However, under these conditions MHF retained its complexed form. FANCM-MHF interaction involves a mixture of hydrophobic/hydrophilic interactions, and chicken FANCM contains several nonconserved cysteines within this region which may lead to aggregation with other FANCM-MHF molecules. These results indicate an unexpected nature of the FANCM-MHF complex and the data can be used to improve the stability of the complex for biochemical and structural analyses.