A Structure-Based Mechanism for DNA Entry into the Cohesin Ring.

Despite key roles in sister chromatid cohesion and chromosome organization, the mechanism by which cohesin rings are loaded onto DNA is still unknown. Here we combine biochemical approaches and cryoelectron microscopy (cryo-EM) to visualize a cohesin loading intermediate in which DNA is locked between two gates that lead into the cohesin ring. Building on this structural framework, we design experiments to establish the order of events during cohesin loading. In an initial step, DNA traverses an N-terminal kleisin gate that is first opened upon ATP binding and then closed as the cohesin loader locks the DNA against the ATPase gate. ATP hydrolysis will lead to ATPase gate opening to complete DNA entry. Whether DNA loading is successful or results in loop extrusion might be dictated by a conserved kleisin N-terminal tail that guides the DNA through the kleisin gate. Our results establish the molecular basis for cohesin loading onto DNA.

Determining the Nutrient Content of Hydroponically-Cultivated Microgreens with Immersible Silicon Photonic Sensors: A Preliminary Feasibility Study.

Microgreens have gained attention for their exceptional culinary characteristics and high nutritional value. The present study focused on a novel approach for investigating the easy extraction of plant samples and the utilization of immersible silicon photonic sensors to determine, on the spot, the nutrient content of microgreens and their optimum time of harvest. For the first time, it was examined how these novel sensors can capture time-shifting spectra caused by the molecules' dynamic adhesion onto the sensor surface. The experiment involved four types of microgreens (three types of basil and broccoli) grown in a do-it-yourself hydroponic installation. The sensors successfully distinguished between different plant types, showcasing their discriminative capabilities. To determine the optimum harvest time, this study compared the sensor data with results obtained through standard analytical methods. Specifically, the total phenolic content and antioxidant activity of two basil varieties were juxtaposed with the sensor data, and this study concluded that the ideal harvest time for basil microgreens was 14 days after planting. This finding highlights the potential of the immersible silicon photonic sensors for potentially replacing time-consuming analytical techniques. By concentrating on obtaining plant extracts, capturing time-shifting spectra, and assessing sensor reusability, this research paves the way for future advancements in urban farming.

Toughening Effect of 2,5-Furandicaboxylate Polyesters on Polylactide-Based Renewable Fibers.

This work presents the successful preparation and characterization of polylactide/poly(propylene 2,5-furandicarboxylate) (PLA/PPF) and polylactide/poly(butylene 2,5-furandicarboxylate) (PLA/PBF) blends in form of bulk and fiber samples and investigates the influence of poly(alkylene furanoate) (PAF) concentration (0 to 20 wt%) and compatibilization on the physical, thermal, and mechanical properties. Both blend types, although immiscible, are successfully compatibilized by Joncryl (J), which improves the interfacial adhesion and reduces the size of PPF and PBF domains. Mechanical tests on bulk samples show that only PBF is able to effectively toughen PLA, as PLA/PBF blends with 5-10 wt% PBF showed a distinct yield point, remarkable necking propagation, and increased strain at break (up to 55%), while PPF did not show significant plasticizing effects. The toughening ability of PBF is attributed to its lower glass transition temperature and greater toughness than PPF. For fiber samples, increasing the PPF and PBF amount improves the elastic modulus and mechanical strength, particularly for PBF-containing fibers collected at higher take-up speeds. Remarkably, in fiber samples, plasticizing effects are observed for both PPF and PBF, with significantly higher strain at break values compared to neat PLA (up to 455%), likely due to a further microstructural homogenization, enhanced compatibility, and load transfer between PLA and PAF phases following the fiber spinning process. SEM analysis confirms the deformation of PPF domains, which is probably due to a "plastic-rubber" transition during tensile testing. The orientation and possible crystallization of PPF and PBF domains contribute to increased tensile strength and elastic modulus. This work showcases the potential of PPF and PBF in tailoring the thermo-mechanical properties of PLA in both bulk and fiber forms, expanding their applications in the packaging and textile industry.

A Punctuated Equilibrium Model for Conceptual Change and the Role of Neo-Piagetian Constructs: Methodology, Empirical Evidence, and Theoretical Perspectives.

This study explores the hypothesis related to a punctuated equilibrium process for conceptual change in science learning, in conjunction with the effects of four cognitive variables: logical thinking and field-dependence/field-independence, divergent and convergent thinking. The participants were fifth and sixth-grade elementary school pupils involved in different tasks, who were asked to describe and interpret chemical phenomena. Latent Class Analysis (LCA) was applied to children's responses, and three clusters or latent classes (LC1, LC2, and LC3) were identified, corresponding to hierarchical levels of conceptual understanding. The ensued LCs align with the theoretical conjecture about a stepwise conceptual change process that might go through various stages or mental models. These levels or stages are conceptualized as attractors, and changes between them were modeled as cusp catastrophes using the four cognitive variables as controls. The analysis showed that logical thinking acted as asymmetry factor, while field-dependence/field-independence, divergent and convergent thinking acted as bifurcation variables. This analytic approach presents a methodology for investigating conceptual change as a punctuated equilibrium process that adds to the nonlinear dynamical research with important implications for theories of conceptual change in science education and psychology as well. Discussion on the new perspective embracing the meta-theoretical framework of complex adaptive systems (CDS) is provided.

Can thromboprophylaxis build a link for cancer patients undergoing surgical and/or chemotherapy treatment? The MeTHOS cohort study.

BACKGROUND: Patients with active cancer have a 4-sevenfold increased risk for venous thromboembolism (VTE) especially during systematic anticancer treatment. Simultaneously, surgery is an additional risk factor. METHODS: The Metaxas's Hospital THromboprophylaxis program in Oncological & Surgical Patients (MeTHOS) is a prospective, phase IV, observational, non-interventional cohort study, aiming to record the thromboprophylaxis practice patterns in high-risk active cancer patients undergoing surgical and/or chemotherapy treatment. RESULTS: We are reporting results from 291 ambulatory patients (median age: 67 years, Q1-Q3: 59-73 years, 54.6% males) who received anti-neoplastic treatment and administered thromboprophylaxis. 59.8% had cardiovascular disease (mostly hypertension), 76.6% were reported as having at least one comorbidity, while 27.5% and 15.8% accumulated two and three comorbidities, respectively. 94.9% of the patients were receiving highly thrombogenic agents such as platinum-based agents, 5-FU, immunotherapy, antiangiogenics/anti-VEGF, or erythropoietin. 26.5% of the patients were initially surgically treated. In terms of anticoagulation, all patients were treated with tinzaparin (fixed dose, 10,000 Anti-Xa IU, OD). The median anticoagulation duration was 6.2 months. Six thrombotic events were observed (2.06%, 95% CI: 0.76-4.43%): 5 were DVT, and one PE. With respect to safety, 7 bleeding events occurred (2.6%, 95% CI: 1.0-5.3%); 6 of them were minor. CONCLUSIONS: Thromboprophylaxis with LMWH in patients with active cancer and high thrombotic burden was safe and effective. Intermediate dose of tinzaparin seems to be an appropriate agent for cancer-associated thromboprophylaxis management. CLINICAL TRIAL REGISTRATION: ClinicalTrials.gov: NCT04248348.

Identifying SARS-CoV-2 antiviral compounds by screening for small molecule inhibitors of Nsp5 main protease.

The coronavirus 2019 (COVID-19) pandemic, caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), spread around the world with unprecedented health and socio-economic effects for the global population. While different vaccines are now being made available, very few antiviral drugs have been approved. The main viral protease (nsp5) of SARS-CoV-2 provides an excellent target for antivirals, due to its essential and conserved function in the viral replication cycle. We have expressed, purified and developed assays for nsp5 protease activity. We screened the nsp5 protease against a custom chemical library of over 5000 characterised pharmaceuticals. We identified calpain inhibitor I and three different peptidyl fluoromethylketones (FMK) as inhibitors of nsp5 activity in vitro, with IC50 values in the low micromolar range. By altering the sequence of our peptidomimetic FMK inhibitors to better mimic the substrate sequence of nsp5, we generated an inhibitor with a subnanomolar IC50. Calpain inhibitor I inhibited viral infection in monkey-derived Vero E6 cells, with an EC50 in the low micromolar range. The most potent and commercially available peptidyl-FMK compound inhibited viral growth in Vero E6 cells to some extent, while our custom peptidyl FMK inhibitor offered a marked antiviral improvement.

Synthesis and Photolytic Assessment of Nitroindolinyl-Caged Calcium Ion Chelators.

Neuroactive amino acids derivatised at their carboxylate groups with a photolabile nitroindolinyl group are highly effective reagents for the sub-µs release of neuroactive amino acids in physiological solutions. However, the same does not apply in the case of calcium ion chelators. In this study, nitroindolinyl-caged BAPTA is found to be completely photostable, whereas nitroindolinyl-caged EDTA photolyses only when saturated with calcium ions.

Author Correction: MCT2 mediates concentration-dependent inhibition of glutamine metabolism by MOG.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Nonlinear Dynamic Effects of Convergent and Divergent Thinking in the Conceptual Change Process: Empirical Evidence from Primary Education.

The present study explores the role of convergent and divergent thinking in learning sciences from the nonlinear dynamical system (NDS) perspective. The participants (N=375) were fifth and sixth graders, aged 11-12, who were taking an introductory course in science. Students' understanding of physical phenomena, such as melting, boiling and evaporation was investigated as a function of four neo-Piagetian constructs via the difference-equation cusp catastrophe model. The nonlinear models where logical thinking acted as the asymmetry factor and field dependence/ independence, convergent thinking and divergent thinking acted as bifurcation factors, were superior, explaining 43-44% of the variance, whereas their linear alternatives explained 0-18%. Empirical evidence regarding the role of the above neo-Piagetian constructs at these early ages is reported for the first time and contributes to theory development within the NDS framework. Further, discussion about the significance of the findings is provided.

Synechococcus elongatus PCC7942: a cyanobacterium cell factory for producing useful chemicals and fuels under abiotic stress conditions.

Sucrose, a compatible osmolyte in cyanobacteria, functions both as an energy reserve and as osmoprotectant. Sugars are the most common substrates used by microorganisms to produce hydrogen (H2) by means of anaerobic dark fermentation. Cells of the unicellular, non-nitrogen fixing, freshwater cyanobacterium Synechococcus elongatus PCC7942 accumulate sucrose under salt stress. In the present work, we used this cyanobacterium and a genetically engineered strain of it (known as PAMCOD) to investigate the optimal conditions for (a) photosynthetic activity, (b) cell proliferation and (c) sucrose accumulation, which are necessary for H2 production via anaerobic dark fermentation of the accumulated sucrose. PAMCOD (Deshnium et al. in Plant Mol Biol 29:897-902, 1995) contains the gene codA that codes for choline oxidase, the enzyme which converts choline to the zwitterion glycine betaine. Glycine betaine is a compatible osmolyte which increases the salt tolerance of Synechococcus elongatus PCC7942. Furthermore, glycine betaine maintains cell proliferation under salt stress and results in increased sucrose accumulation. In the present study, we examine the environmental factors, such as the NaCl concentration, the culture medium pH, and the carbon dioxide content of the air bubbled through it. At optimal conditions, sucrose accumulated in the cyanobacteria cells up to 13.5 mol per mole Chl a. Overall, genetically engineered Synechococcus elongatus PCC7942 produces sucrose in sufficient quantities such that it may be a viable alternative (a) to sucrose synthesis, and (b) to H2 formation via anaerobic dark fermentation.

MCT2 mediates concentration-dependent inhibition of glutamine metabolism by MOG.

α-Ketoglutarate (αKG) is a key node in many important metabolic pathways. The αKG analog N-oxalylglycine (NOG) and its cell-permeable prodrug dimethyloxalylglycine (DMOG) are extensively used to inhibit αKG-dependent dioxygenases. However, whether NOG interference with other αKG-dependent processes contributes to its mode of action remains poorly understood. Here we show that, in aqueous solutions, DMOG is rapidly hydrolyzed, yielding methyloxalylglycine (MOG). MOG elicits cytotoxicity in a manner that depends on its transport by monocarboxylate transporter 2 (MCT2) and is associated with decreased glutamine-derived tricarboxylic acid-cycle flux, suppressed mitochondrial respiration and decreased ATP production. MCT2-facilitated entry of MOG into cells leads to sufficiently high concentrations of NOG to inhibit multiple enzymes in glutamine metabolism, including glutamate dehydrogenase. These findings reveal that MCT2 dictates the mode of action of NOG by determining its intracellular concentration and have important implications for the use of (D)MOG in studying αKG-dependent signaling and metabolism.

Scaffold-hopping identifies furano[2,3-d]pyrimidine amides as potent Notum inhibitors.

The carboxylesterase Notum is a key negative regulator of the Wnt signaling pathway by mediating the depalmitoleoylation of Wnt proteins. Our objective was to discover potent small molecule inhibitors of Notum suitable for exploring the regulation of Wnt signaling in the central nervous system. Scaffold-hopping from thienopyrimidine acids 1 and 2, supported by X-ray structure determination, identified 3-methylimidazolin-4-one amides 20-24 as potent inhibitors of Notum with activity across three orthogonal assay formats (biochemical, extra-cellular, occupancy). A preferred example 24 demonstrated good stability in mouse microsomes and plasma, and cell permeability in the MDCK-MDR1 assay albeit with modest P-gp mediated efflux. Pharmacokinetic studies with 24 were performed in vivo in mouse with single oral administration of 24 showing good plasma exposure and reasonable CNS penetration. We propose that 24 is a new chemical tool suitable for cellular studies to explore the fundamental biology of Notum.

Eugene I. Rabinowitch: A prophet of photosynthesis and of peace in the world.

More than 45 years have passed since Eugene I. Rabinowitch died, on May 15, 1973, at the age of 75, but many still remember him as a photosynthesis giant, the author of a 2000-page "Bible" on photosynthesis, a great chemist and physicist, a discoverer of several basic photoreactions, one of the founders of modern biophysics, a peacemaker, a poet, an architect, an artist, a wonderful human being, and above all a great mentor. Sir John Rotblatt cited Eugene Rabinowitch, together with Bertrand Russell, for their key contributions that led to the Nobel Peace Prize awarded in 1995 jointly to Rotblatt and the Pugwash Conferences on Science and World Affairs "for their efforts to diminish the part played by nuclear arms in international politics and, in the longer run, to eliminate such arms." Already in 1965, Eugene Rabinowitch had received the prestigious Kalinga Prize from UNESCO "in recognition of his work to encourage international cooperation among scientists and to bring to light the potential dangers of science to the public."

Kinetics of Crystallization and Thermal Degradation of an Isotactic Polypropylene Matrix Reinforced with Graphene/Glass-Fiber Filler.

Polypropylene composites reinforced with a filler mixture of graphene nanoplatelet-glass fiber were prepared by melt mixing, while conventional composites containing graphene nanoplatelet and glass fiber were prepared for comparative reasons. An extensive study of thermally stimulated processes such as crystallization, nucleation, and kinetics was carried out using Differential Scanning Calorimetry and Thermogravimetric Analysis. Moreover, effective activation energy and kinetic parameters of the thermal decomposition process were determined by applying Friedman's isoconversional differential method and multivariate non-linear regression method. It was found that the graphene nanoplatelets act positively towards the increase in crystallization rate and nucleation phenomena under isothermal conditions due to their large surface area, inherent nucleation activity, and high filler content. Concerning the thermal degradation kinetics of polypropylene graphene nanoplatelets/glass fibers composites, a change in the decomposition mechanism of the matrix was found due to the presence of graphene nanoplatelets. The effect of graphene nanoplatelets dominates that of the glass fibers, leading to an overall improvement in performance.

Thermal Decomposition Kinetics and Mechanism of In-Situ Prepared Bio-based Poly(propylene 2,5-furan dicarboxylate)/Graphene Nanocomposites.

Bio-based polyesters are a new class of materials that are expected to replace their fossil-based homologues in the near future. In this work, poly(propylene 2,5-furandicarboxylate) (PPF) nanocomposites with graphene nanoplatelets were prepared via the in-situ melt polycondensation method. The chemical structure of the resulting polymers was confirmed by 1H-NMR spectroscopy. Thermal stability, decomposition kinetics and the decomposition mechanism of the PPF nanocomposites were studied in detail. According to thermogravimetric analysis results, graphene nanoplatelets did nοt affect the thermal stability of PPF at levels of 0.5, 1.0 and 2.5 wt.%, but caused a slight increase in the activation energy values. Pyrolysis combined with gas chromatography and mass spectroscopy revealed that the decomposition mechanism of the polymer was not altered by the presence of graphene nanoplatelets but the extent of secondary homolytic degradation reactions was increased.

An improved, scalable synthesis of Notum inhibitor LP-922056 using 1-chloro-1,2-benziodoxol-3-one as a superior electrophilic chlorinating agent.

Background: The carboxylesterase Notum has been shown to act as a key negative regulator of the Wnt signalling pathway by mediating the depalmitoleoylation of Wnt proteins. LP-922056 (1) is an orally active inhibitor of Notum. We are investigating the role of Notum in modulating Wnt signalling in the central nervous system and wished to establish if 1 would serve as a peripherally restricted control. An accessible and improved synthetic route would allow 1 to become more readily available as a chemical tool to explore the fundamental biology of Notum and build target validation to underpin new drug discovery programs. Results: An improved, scalable synthesis of 1 is reported. Key modifications include: (1) the introduction of the C7-cyclopropyl group was most effectively achieved with a Suzuki-Miyaura cross-coupling reaction with MIDA-boronate 11 (5 → 6), and (2) C6 chlorination was performed with 1-chloro-1,2-benziodoxol-3-one (12) (6 → 7) as a mild and selective electrophilic chlorination agent. This 7-step route from 16 has been reliably performed on large scale to produce multigram quantities of 1 in good efficiency and high purity. Pharmacokinetic studies in mouse showed CNS penetration of 1 is very low with a brain/plasma concentration ratio of just 0.01. A small library of amides 17 were prepared from acid 1 to explore if 1 could be modified to deliver a CNS penetrant tool by capping off the acid as an amide. Although significant Notum inhibition activity could be achieved, none of these amides demonstrated the required combination of metabolic stability along with cell permeability without evidence of P-gp mediated efflux. Conclusion: Mouse pharmacokinetic studies demonstrate that 1 is unsuitable for use in models of disease where brain penetration is an essential requirement of the compound but would be an ideal peripherally restricted control. These data will contribute to the understanding of drug levels of 1 to overlay with appropriate in vivo efficacy endpoints, i.e., the PK-PD relationship. The identification of a suitable analogue of 1 (or 17) which combines Notum inhibition with CNS penetration would be a valuable chemical probe for investigating the role of Notum in disease models.

The extraordinary longevity of kleptoplasts derived from the Ross Sea haptophyte Phaeocystis antarctica within dinoflagellate host cells relates to the diminished role of the oxygen-evolving Photosystem II and to supplementary light harvesting by mycosporine-like amino acid/s.

The haptophyte Phaeocystis antarctica and the novel Ross Sea dinoflagellate that hosts kleptoplasts derived from P. antarctica (RSD; R.J. Gast et al., 2006, J. Phycol. 42 233-242) were compared for photosynthetic light harvesting and for oxygen evolution activity. Both chloroplasts and kleptoplasts emit chlorophyll a (Chl a) fluorescence peaking at 683nm (F683) at 277K and at 689 (F689) at 77K. Second derivative analysis of the F689 band at 77K revealed two individual contributions centered at 683nm (Fi-683) and at 689 (Fi-689). Using the p-nitrothiophenol (p-NTP) treatment of Kobayashi et al. (Biochim. Biophys. Acta 423 (1976) 80-90) to differentiate between Photosystem (PS) II and I fluorescence emissions, we could identify PS II as the origin of Fi-683 and PS I as the origin of Fi-689. Both emissions could be excited not only by Chl a-selective light (436nm) but also by mycosporine-like amino acids (MAAs)-selective light (345nm). This suggests that a fraction of MAAs must be proximal to Chls a and, therefore, located within the plastids. On the basis of second derivative fluorescence spectra at 77K, of p-NTP resolved fluorescence spectra, as well as of PSII-driven oxygen evolution activities, PS II appears substantially less active (~1/5) in dinoflagellate kleptoplasts than in P. antarctica chloroplasts. We suggest that a diminished role of PS II, a known source of reactive oxygen species, and a diminished dependence on nucleus-encoded light-harvesting proteins, due to supplementary light-harvesting by MAAs, may account for the extraordinary longevity of RSD kleptoplasts.

Frederick Yi-Tung Cho (1939-2011) : His PhD days in Biophysics, the Photosynthesis Lab, and his patents in engineering physics.

We present here a Tribute to Frederick Yi-Tung Cho (1939-2011), an innovative and ingenious biophysicist and an entrepreneur. He was one of the 4 earliest PhD students [see: Cederstrand (1965)-Carl Nelson Cederstrand; coadvisor: Eugene Rabinowitch; Papageorgiou (1968)-George C. Papageorgiou (coauthor of this paper); and Munday (1968)-John C. Munday Jr. (also a coauthor of this paper)] of one of us (Govindjee) in Biophysics at the University of Illinois at Urbana-Champaign (UIUC) during the late 1960s (1963-1968). Fred was best known, in the photosynthesis circle for his pioneering work on low temperature (down to liquid helium temperature, 4 K) absorption and fluorescence spectroscopy of photosynthetic systems; he showed temperature independence of excitation energy transfer from (i) chlorophyll (Chl) b to Chl a and (ii) from Chl a 670 to Chl a 678; and temperature dependence of energy transfer from the phycobilins to Chl a and from Chl a 678 to its suggested trap. After doing research in biophysics of photosynthesis, Fred shifted to do research in solid-state physics/engineering in the Government Electronics Division (Group) of the Motorola Company, Scottsdale, Arizona, from where he published research papers in that area and had several patents granted. We focus mainly on his days at the UIUC in context of the laboratory in which he worked. We also list some of his papers and most of his patents in engineering physics. His friends and colleagues have correctly described him as an innovator and an ingenious scientist of the highest order. On the personal side, he was a very easy-going and amiable individual.

Effects of exogenous ß-carotene, a chemical scavenger of singlet oxygen, on the millisecond rise of chlorophyll a fluorescence of cyanobacterium Synechococcus sp. PCC 7942.

Singlet-excited oxygen (1O 2* ) has been recognized as the most destructive member of the reactive oxygen species (ROS) which are formed during oxygenic photosynthesis by plants, algae, and cyanobacteria. ROS and 1O 2* are known to damage protein and phospholipid structures and to impair photosynthetic electron transport and de novo protein synthesis. Partial protection is afforded to photosynthetic organism by the ß-carotene (ß-Car) molecules which accompany chlorophyll (Chl) a in the pigment-protein complexes of Photosystem II (PS II). In this paper, we studied the effects of exogenously added ß-Car on the initial kinetic rise of Chl a fluorescence (10-1000 µs, the OJ segment) from the unicellular cyanobacterium Synechococcus sp. PCC7942. We show that the added ß-Car enhances Chl a fluorescence when it is excited at an intensity of 3000 µmol photons m-2 s-1 but not when excited at 1000 µmol photons m-2 s-1. Since ß-Car is an efficient scavenger of 1O 2* , as well as a quencher of 3Chl a * (precursor of 1O 2* ), both of which are more abundant at higher excitations, we assume that the higher Chl a fluorescence in its presence signifies a protective effect against photo-oxidative damages of Chl proteins. The protective effect of added ß-Car is not observed in O2-depleted cell suspensions. Lastly, in contrast to ß-Car, a water-insoluble molecule, a water-soluble scavenger of 1O 2* , histidine, provides no protection to Chl proteins during the same time period (10-1000 µs).

A backbone amide protecting group for overcoming difficult sequences and suppressing aspartimide formation.

A backbone amide bond protecting group, 2-hydroxy-4-methoxy-5-nitrobenzyl (Hmnb), improved the synthesis of aggregation and aspartimide-prone peptides. Introduction of Hmnb is automated and carried out during peptide assembly by addition of 4-methoxy-5-nitrosalicylaldehyde to the peptidyl-resin and on-resin reduction to the secondary amine. Acylation of the hindered secondary amine is aided by the formation of an internal nitrophenol ester that undergoes a favourable O,N intramolecular acyl transfer. This activated ester participates in the coupling and generally gives complete reaction with standard coupling conditions. Hmnb is easily available in a single preparative step from commercially available material. Different methods for removing the amide protecting group were explored. The protecting group is labile to acidolysis, following reduction of the nitro group to the aniline. The two main uses of backbone protection of preventing aspartimide formation and of overcoming difficult sequences are demonstrated, first with the synthesis of a challenging aspartimide-prone test sequence and then with the classic difficult sequence ACP (65-74) and a 23-mer homopolymer of polyalanine.