Backbone and Ile-δ1, Leu,Val methyl <Superscript>1</Superscript>H, <Superscript>15</Superscript>N,and <Superscript>13</Superscript>C,chemical shift assignments for <Emphasis Type="Italic">Rhizopus chinensis</Emphasis> lipase

Lipase r27RCL is a 296-residue, 33 kDa monomeric enzyme with high ester hydrolysis activity, which has significant applications in the baking, paper and leather industries. The lipase gene proRCL from Rhizopus microsporus var. chinensis (also Rhizopus chinensis) CCTCC M201021 was cloned as a fusion construct C-terminal to a maltose-binding protein (MBP) tag, and expressed as MBP-proRCL in an Escherichia coli BL21 trxB (DE3) expression system with uniform ²H,¹³C,¹⁵N-enrichment and Ile-δ1, Leu, and Val ¹³CH₃ methyl labeling. The fusion protein was hydrolyzed by Kex2 protease at the recognition site Lys-Arg between residues ?29 and ?28 of the prosequence, producing the enzyme form called r27RCL. Here we report extensive backbone ¹H, ¹⁵N, and ¹³C, as well as Ile-δ1, Leu, and Val side chain methyl, NMR resonance assignments for r27RCL.     

Mechanisms of mechanosensing — mechanosensitive channels,function and re-engineering

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Development,optimization, and biovalidation of <Superscript>99m</Superscript>Tc–insulin complex

Human biosynthetic insulin is a polypeptide hormone that plays an important and essential role in control of the level of carbohydrate, protein, and fat metabolism in the blood. Human pancreatic insulin was labeled with ^99mTc to form a new radiopharmaceutical with a labeling yield of 99 ± 1% under optimum conditions: 0.1 mL insulin, pH 7, 25 μg stannous chloride, 1 mL (19 mCi) of pertechnetate, room temperature, and 10 min reaction time. The ^99mTc–insulin complex was examined using paper chromatography, ITLC, electrophoresis, and HPLC. In addition, in vitro and in vivo study of ^99mTc–insulin complex was performed at different time intervals.     

New κ-PNN′- and κ-PNN′O-polydentate ligands: Synthesis, coordination and structural studies

The three-step synthesis of new mixed P/N/N′/O-donor ligands C₆H₃(OH){2-NHC(O)CH₂NCHC₆H₄PPh₂}(4-CH₃) 3a·HH and C₆H₄(OH){3-NHC(O)CH₂NCHC₆H₄PPh₂} 3b·HH, by Schiff base condensation of the 1° amines C₆H₃(OH){2-NHC(O)CH₂NH₂}(4-CH₃) 2a or C₆H₄(OH){3-NHC(O)CH₂NH₂} 2b with C₆H₄(CHO)(2-PPh₂) in refluxing EtOH, is described. Reaction of 1 equiv. of 3a·HH or 3b·HH with MCl₂(cod) (M = Pt, Pd; cod = cycloocta-1,5-diene) affords the κ²-PN-chelate complexes MCl₂(3a·HH) (M = Pd 4a; M = Pt 4b) and MCl₂(3b·HH) (M = Pt 4c). The dichlorometal(II) complexes 4d and 4e, bearing instead a pendant 4-phenolic group, were similarly prepared (in >90% yield). Chloro-bridge cleavage of [Pd(μ-Cl)(η³-C₃H₅)]₂ with 3a·HH or 3b·HH gave the monocationic κ²-PN-chelate complexes [Pd(η³-C₃H₅)(3a·HH)]Cl 5a or [Pd(η³-C₃H₅)(3b·HH)]Cl 5b, respectively. Elimination of cod, and single CH₃ protonation, from Pt(CH₃)₂(cod) upon reaction with 1 equiv. of 3a·HH or 3b·HH in C₇H₈ at room temperature afforded the neutral complexes C₆H₃(OH){2-NC(O)CH₂NCHC₆H₄PPh₂Pt(CH₃)}(4-CH₃) 6a and C₆H₄(OH){3-NC(O)CH₂NCHC₆H₄PPh₂Pt(CH₃)} 6b, respectively bearing a monoanionic (3a·H⁻ or 3b·H⁻) κ³-PNN′-tridentate ligand. Amide and phenol deprotonation were readily achieved, using KO^tBu as base, to give high yields of the κ⁴-PNN′O-tetradentate complexes C₆H₃(O){2-NC(O)CH₂NCHC₆H₄PPh₂Pd}(4-CH₃) 7a and C₆H₃(O){2-NC(O)CH₂NCHC₆H₄PPh₂Pt}(4-CH₃) 7b bearing the dianionic ligand 3a²⁻. All new compounds have been characterised by multinuclear NMR, FTIR, mass spectroscopy and microanalysis. Single crystal X-ray studies have been performed on compounds 1b·1.5CH₂Cl₂, 3b·HH·0.5Et₂O, 6b·CHCl₃ and 7b·0.5Et₂O.     

Electric-field-induced spin switch of endohedral dodecahedrane heterodimers H@C<Subscript>20</Subscript>H<Subscript>n</Subscript>–C<Subscript>20</Subscript>H<Subscript>n</Subscript>@M (M= Cu,Ag and Au,n = 15, 18, and 19): a theoretical study

We designed nine endohedral dodecahedrane heterodimers H@C₂₀H_n-C₂₀H_n@M (M = Cu, Ag, and Au, n = 15, 18, and 19) that may act as single-molecule spin switches, and we predicted theoretically that the ground states of the dimmers shift from low-spin states (S = 0) to the high-spin states (S = 1) under an external electric field applied parallel or perpendicular to the molecular symmetry axes, consisting well with the analyses of Stark effect. Molecular orbitals analyses provide an intuitive insight into the spin crossover behavior. This study expands the application of endohedral chemistry and provides new molecules for designing single-molecule spin switch.     

Effects of activation of μ-, κ<Subscript>1</Subscript>-, δ<Subscript>1</Subscript>-, δ<Subscript>2</Subscript>-, and ORL<Subscript>1</Subscript>-receptors on heart resistance to the pathogenic action of delayed ischemia and reperfusion

Different subtypes of opioid receptors (OR) were activated in rats in vivo to study the activation effect on the heart’s resistance to ischemia and reperfusion. It has been established that administration of deltorphin II, a selective δ₂-OR agonist, lowered the infarct size/area at risk index (IS/AAR) by 23%. Naltrexone, naloxone methiodide (an OR inhibitor not penetrating the blood-brain barrier (BBB)), and naltriben (δ₂-antagonist) eliminated the cardioprotective effect of deltorphin II, while BNTX (a δ₁-antagonist) produced no effect on the cardioprotective action of the δ₂-agonist. The infarct-reducing effect of deltorphin II was eliminated by administration of chelerythrine (a protein kinase C (PKC) inhibitor), glibenclamide (a K_ATP-channels inhibitor), and 5-hydroxydecanoate (a mitochondrial K_ATP-channel blocker). Administration of other opioids did not reduce the IS/AAR index. It has been established that all the deltorphins manifest antiarrhythmic potency. Other opioids do not produce any effect on the incidence of arrhythmia occurrences. The antiarrhythmic effect of deltorphin II was eliminated by preliminary administration of naltrexone, naloxone methiodide, and naltriben, but BNTX did not affect the δ₂-agonist’s anti-arrhythmic effect. The preliminary administration of chelerythrine, a PKC inhibitor, eliminated the δ₂ agonist’s antiarrhythmic action. However, glibenclamide and 5-hydroxydecanoate did not alter the antiarrhythmic effect by deltorphin II. Therefore, activation of the peripheral δ₂-ORs reduces the infarct size and prevents the onset of arrhythmias. The antiarrhythmic effect of the δ₂-OR stimulation is mediated by activating PKC and opening the mitochondrial K_ATP-channels. PKC participates in the antiarrhythmic effect of the δ₂-OR activation, but this effect does not depend on the condition of K_ATP-channels.     

Influence of Acute and Chronic Treadmill Exercise on Rat Plasma Lactate and Brain NPY,L-ENK,DYN A<Subscript>1–13</Subscript>

SUMMARY This study was designed to investigate the effect of acute and chronic high-intensity treadmill exercise on changes in plasma lactate and brain neuropeptide (NPY), leucine-enkephalin (L-ENK), and dynorpin A_1–13 (DYN A_1–13). Avidin–biotin complex (ABC) immunohistochemistry and image pattern analysis were used to observe the effect of chronic (total 7 weeks) and acute treadmill exercise (an initial speed of 15 m min⁻¹ gradually increased to 35 m min⁻¹ with 0°, 20–25 min per day duration) on the changes of NPY, L-ENK, and DYN A_1–13 in different areas of rat brain. Plasma lactate was also measured in response to such exercise. Compared with preexercise control (P < 0.01), plasma lactate concentration significantly increased in the immediate postexercise; but it returned to the normal level soon after the 30 min postexercise. The content of NPY in paraventricular (PVN), dorsomedial (DMN), and ventromedial (VMN) hypothalamic nuclei continued to increase in 0, 30, and 180 min postexercise compared with preexercise control (P < 0.01). The content of L-ENK in caudate-putamen (CPu) significantly increased in the immediate postexercise compared with preexercise control (P < 0.01), but it gradually returned to the normal level after the 180 min postexercise. However, the content of DYN A_1–13 in PVN rose substantially only in 30 min postexercise in comparison with the preexercise control (P < 0.01). Thus, different changes of NPY, L-ENK, and DYN A_1–13 in response to such high-intensity exercise depend on the brain region and the time examined, especially, the contents of NPY in different brain regions continuously remain at a high level after such high-intensity exercise. And this high level might reduce energy expenditure and thus contribute to the stimulation of brain NPY neurons.     

MP2, density functional theory, and molecular mechanical calculations of C-H···π and hydrogen bond interactions in a cellulose-binding module-cellulose model system

Exploring non-covalent interactions, such as C-H···π stacking and classical hydrogen bonding (H-bonding), between carbohydrates and carbohydrate-binding modules (CBMs) is an important task in glycobiology. The present study focuses on intermolecular interactions, such as C-H?π (sugar-aromatic stacking) and H-bonds, between methyl β-d-glucopyranoside and l-tyrosine—a proxy model system for a cellulose-CBM complex. This work has made use of various types of quantum mechanics (QM) and molecular mechanics (MM) methods to determine which is the most accurate and computationally efficient. The calculated interaction potential energies ranged between −24 and −38 kJ/mol. The larger interaction energy is due to H-bonding between the phenyl hydroxyl of tyrosine and the O4 of the sugar. Density functional theory (DFT) methods, such as BHandHLYP and B3LYP, exaggerate the H-bond. Although one of the MM methods (viz. MM+) considered in this study does maintain the C-H?π stacking configuration, it underestimates the interaction energy due to the loss of the H-bond. When the O-H bond vector is in the vicinity of O4 (O-H?O4 ≈ 2 Å, e.g., in the case of MP2/6-31G(d)), the torsional energy drops to a minimum. For this configuration, natural bond orbital (NBO) analysis also supports the presence of this H-bond which arises due to orbital interaction between one lone pair of the sugar O4 and the σ∗(O-H) orbital of the phenyl group of tyrosine. The stabilization energy due to orbital delocalization of the H-bonded system is ∼13 kJ/mol. This H-bond interaction plays an important role in controlling the CH/π interaction geometry. Therefore, the C-H?π dispersive interaction is the secondary force, which supports the stabilization of the complex. The meta-hybrid DFT method, M05-2X, with the 6-311++G(d,p) basis set agrees well with the MP2 results and is less computationally expensive. However, the M05-2X method is strongly basis set dependent in describing this CH/π interaction. Computed IR spectra with the MP2/6-31G(d) method show blue shifts for C1-H, C3-H, and C5-H stretching frequencies due to the C-H?π interaction. However, the M05-2X/6-311++G(d,p) method shows a small red shift for the C1-H stretching region and blue shifts for the C2-H and C3-H stretches. For the aromatic tyrosine C_δ1-C_ε1 and C_δ2-C_ε2 bonds in the complex, the calculated IR spectra show red shifts of 12 cm⁻¹ (MP2/6-31G(d)) and 5 cm⁻¹ (M05-2X/6-311++G(d,p)). This study also reports the upfield shifts of computed ¹H NMR chemical shifts due to the C-H?π interaction.     

Interactions between oestrogen and 1α,25(OH)<Subscript>2</Subscript>-vitamin D<Subscript>3</Subscript> signalling and their roles in spermatogenesis and spermatozoa functions

Oestrogens and 1α,25(OH)₂-vitamin D₃ (1,25-D₃) are steroids that can provide effects by binding to their receptors localised in the cytoplasm and in the nucleus or the plasma membrane respectively inducing genomic and non-genomic effects. As confirmed notably by invalidation of the genes, coding for their receptors as tested with mice with in vivo and in vitro treatments, oestrogens and 1,25-D₃ are regulators of spermatogenesis. Moreover, some functions of ejaculated spermatozoa as viability, DNA integrity, motility, capacitation, acrosome reaction and fertilizing ability are targets for these hormones. The studies conducted on their mechanisms of action, even though not completely elicited, have allowed the demonstration of putative interactions between their signalling pathways that are worth examining more closely. The present review focuses on the elements regulated by oestrogens and 1,25-D₃ in the testis and spermatozoa as well as the interactions between the signalling pathways of both hormones.     

Perovskite Solar Cells: Can We Go Organic‐Free,Lead‐Free,and Dopant‐Free?

Having demonstrated incredibly fast progress in power conversion efficiency, rising to a level comparable with that of crystalline silicon cells, lead‐based organic–inorganic hybrid perovskite solar cells are now facing the stability tests needed for industrialization. Poor thermal stability (<150 °C) owing to organic constituents and interlayer diffusion of materials (dopants), and environmental incompatibility due to Pb has surged the development of organic‐free, Pb‐free perovskites and dopant‐free hole transport materials (HTMs). The recent rapid increase in efficiency of cells based on inorganic perovskites, crossing 18%, demonstrates the great potential of inorganic perovskites as thermally stable and high‐efficiency cells. Although all kinds of Pb‐free perovskites lag in efficiency in comparison to the hybrid and inorganic perovskites, they also demonstrate better structural and environmental stability. The performance of dopant‐free HTMs matching/surpassing dopant‐containing HTMs makes the former a better choice for stability. Even though the efforts to enhance the stability of Pb‐based hybrid perovskites should continue by different techniques, organic‐free and lead‐free perovskites, and dopant‐free HTMs must be pursued with greater interest for the future. This review describes the present issues and possible strategies to address them, and thus will help to improve the overall performance of robust organic‐free, Pb‐free, and dopant‐free perovskite solar cells.     

Which practices co‐deliver food security,climate change mitigation and adaptation,and combat land degradation and desertification?

There is a clear need for transformative change in the land management and food production sectors to address the global land challenges of climate change mitigation, climate change adaptation, combatting land degradation and desertification, and delivering food security (referred to hereafter as “land challenges”). We assess the potential for 40 practices to address these land challenges and find that: Nine options deliver medium to large benefits for all four land challenges. A further two options have no global estimates for adaptation, but have medium to large benefits for all other land challenges. Five options have large mitigation potential (>3 Gt CO₂eq/year) without adverse impacts on the other land challenges. Five options have moderate mitigation potential, with no adverse impacts on the other land challenges. Sixteen practices have large adaptation potential (>25 million people benefit), without adverse side effects on other land challenges. Most practices can be applied without competing for available land. However, seven options could result in competition for land. A large number of practices do not require dedicated land, including several land management options, all value chain options, and all risk management options. Four options could greatly increase competition for land if applied at a large scale, though the impact is scale and context specific, highlighting the need for safeguards to ensure that expansion of land for mitigation does not impact natural systems and food security. A number of practices, such as increased food productivity, dietary change and reduced food loss and waste, can reduce demand for land conversion, thereby potentially freeing‐up land and creating opportunities for enhanced implementation of other practices, making them important components of portfolios of practices to address the combined land challenges.     

The interaction of CCl<Subscript>4</Subscript> with Ng (Ng = He,Ne, Ar), O<Subscript>2</Subscript>, D<Subscript>2</Subscript>O and ND<Subscript>3</Subscript>: rovibrational energies,spectroscopic constants and theoretical calculations

This investigation generated rovibrational energies and spectroscopic constants for systems of CCl₄ with Ng (Ng?=?He, Ne, Ar), O₂, D₂O and ND₃ from scattering experimental data, and the results presented are of interest for microwave spectroscopy studies of small halogenated molecules. The rovibrational spectra were obtained through two different approaches (Dunham and DVR) within the improved Lennard Jones (ILJ) model. Spectra were also generated within ordinary Lennard Jones and deviations suggest that the ILJ model should be preferred due to interactions beyond dispersion forces presented in these systems. Data from the literature and additional high level quantum mechanical calculations presented in this work show that these systems should not be considered as van der Waals complexes due to halogen bonding (HB) interactions, and this is especially true for the CCl₄–D₂O and CCl₄–ND₃ complexes. The charge displacement from the latter systems are one order of magnitude higher than the values from literature for CCl₄ and He, Ne, Ar and O₂ systems, and show significant deviations between DFT and Hartree-Fock values not previously reported in the literature.     

Electronic and optical properties of C<Subscript>24</Subscript>, C<Subscript>12</Subscript>X<Subscript>6</Subscript>Y<Subscript>6</Subscript>, and X<Subscript>12</Subscript>Y<Subscript>12</Subscript> (X = B,Al and Y = N,P)

Utilizing first-principles calculations, we studied the electronic and optical properties of C₂₄, C₁₂X₆Y₆, and X₁₂Y₁₂ fullerenes (X?=?B, Al; Y?=?N, P). These fullerenes are energetically stable, as demonstrated by their negative cohesive energies. The energy gap of C₂₄ may be tuned by doping, and the B₁₂N₁₂ fullerene was found to have the largest energy gap. All of the fullerenes had finite optical gaps, suggesting that they are optical semiconductors, and they strongly absorb UV radiation, so they could be used in UV light protection devices. They could also be used in solar cells and LEDs due to their low reflectivities.

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Graphical abstract Possible applications of doped C₂₄ fullerene

     

Duplication and transposition of the<Emphasis Type="Italic"> NF1</Emphasis> pseudogene regions on chromosomes 2, 14, and 22

Numerous NF1 pseudogenes have been identified in the human genome. Those in 2q21, 14q11, and 22q11 form a subset with a similar genomic organization and a high sequence homology. We have studied, by polymerase chain reaction and fluorescence in situ hybridization, the extent of homology of the regions surrounding these NF1 pseudogenes. Our analyses have demonstrated that a fragment of at least 640 kb is homologous between the three regions. Based on previous studies and these new findings, we propose a model for the spreading of the NF1 pseudogene-containing regions. A fragment of approximately 640 kb was first duplicated in chromosome region 2q21 and transposed to 14q11. Subsequently, this fragment was duplicated in 14q11 and transposed to 22q11. A part of the 640-kb fragment in 14q11, with a length of about 430 kb, was further duplicated to a variable extent in 14q11. In addition, we have identified sequences that may facilitate the duplication and transposition of the 640-kb and 430-kb fragments.     

Brain,antibiotics, and microbiota – how do they interplay?

The microbiome and its cross‐talk with the brain have drawn increasing attention lately, since imbalances in the gut microbiota's composition may result in pathogenic dysfunctions affecting brain functioning up to development of neurodegenerative and mental diseases. The current Editorial discusses a study by Gao and coworkers in the current issue of the Journal of Neurochemistry in which the authors use a model of antibiotic‐induced dysbiosis ‐ targeted infusion of antibiotics into the gut ‐ to assess if microbiotic metabolites exert effects on local neurotransmitter expression or contribute to the gut‐brain axis. The authors mechanistically link distal ileal infusion of antibiotics with a change in the levels of microbial metabolites that affect the expression of neurotransmitters in the brain and thereby can participate in the fine‐tuning of the hypothalamic functions, including regulation of visceral and neuroendocrine processes, stress responses, mood and anxiety. Their study thus represents an important step towards our understanding of the brain‐gut axis, with the potential to advance therapeutics.

     

Identification,cloning, and characterization of β-glucosidase from <Emphasis Type="Italic">Ustilago esculenta</Emphasis>

Hydrolytic enzymes responsible for laminarin degradation were found to be secreted during growth of Ustilago esculenta on laminarin. An enzyme involved in laminarin degradation was purified by assaying release of glucose from laminaribiose. Ion-exchange chromatography of the culture filtrate followed by size-exclusion chromatography yielded a 110-kDa protein associated with laminaribiose hydrolysis. LC/MS/MS analysis of the 110-kDa protein identified three peptide sequences that shared significant similarity with a putative glucoside hydrolase family (GH) 3 β-glucosidase in Ustilago maydis. Based on the DNA sequence of the U. maydis GH3 β-glucosidase, a gene encoding a putative GH3 β-glucosidase in U. esculenta (Uebgl3A) was cloned by PCR. Based on the deduced amino acid sequence, the protein encoded by Uebgl3A has a molecular mass of 91 kDa and shares 90% identity with U. maydis GH3 β-glucosidase. Recombinant UeBgl3A expressed in Aspergillus oryzae released glucose from β-1,3-, β-1,4-, and β-1,6-linked oligosaccharides, and from 1,3-1,4-β-glucan and laminarin polysaccharides, indicating that UeBgl3A is a β-glucosidase. Kinetic analysis showed that UeBgl3A preferentially hydrolyzed laminaritriose and laminaritetraose. These results suggest that UeBgl3A is a key enzyme that produces glucose from laminarioligosaccharides during growth of U. esculenta on laminarin.