Purification of low-abundance lysozyme in egg white via free-flow electrophoresis with gel-filtration chromatography

As an effective separation tool, free-flow electrophoresis has not been used for purification of low-abundance protein in complex sample matrix. Herein, lysozyme in complex egg white matrix was chosen as the model protein for demonstrating the purification of low-content peptide via an FFE coupled with gel fitration chromatography (GFC). The crude lysozyme in egg while was first separated via free-flow zone electrophoresis (FFZE). After that, the fractions with lysozyme activity were condensed via lyophilization. Thereafter, the condensed fractions were further purified via a GFC of Sephadex G50. In all of the experiments, a special poly(acrylamide- co-acrylic acid) (P(AM-co-AA)) gel electrophoresis and a mass spectrometry were used for identification of lysozyme. The conditions of FFZE were optimized as follows: 130 μL/min sample flow rate, 4.9 mL/min background buffer of 20 mM pH 5.5 Tris-Acetic acid, 350 V, and 14 °C as well as 2 mg/mL protein content of crude sample. It was found that the purified lysozyme had the purity of 80% and high activity as compared with its crude sample with only 1.4% content and undetectable activity. The recoveries in the first and second separative steps were 65% and 82%, respectively, and the total recovery was about 53.3%. The reasons of low recovery might be induced by diffusion of lysozyme out off P(AM-co-AA) gel and co-removing of high-abundance egg ovalbumin. All these results indicated FFE could be used as alternative tool for purification of target solute with low abundance.     

Synergistic N-Heterocyclic Carbene/Palladium-Catalyzed Umpolung 1,4-Addition of Aryl Iodides to Enals

An umpolung 1,4-addition of aryl iodides to enals promoted by cooperative (terpy)Pd/NHC catalysis was developed that generates various bioactive β,β-diaryl propanoate derivatives. This system is not only the first reported palladium-catalyzed arylation of NHC-bound homoenolates but also expands the scope of NHC-induced umpolung transformations. A diverse array of functional groups such as esters, nitriles, alcohols, and heterocycles are tolerated under the mild conditions. This method also circumvents the use of moisture-sensitive organometallic reagents.     

3-O-Sulfation of Heparan Sulfate Enhances Tau Interaction and Cellular Uptake

Prion-like transcellular spreading of tau in Alzheimer's Disease (AD) is mediated by tau binding to cell surface heparan sulfate (HS). However, the structural determinants for tau–HS interaction are not well understood. Microarray and SPR assays of structurally defined HS oligosaccharides show that a rare 3-O-sulfation (3-O-S) of HS significantly enhances tau binding. In Hs3st1^−/− (HS 3-O-sulfotransferase-1 knockout) cells, reduced 3-O-S levels of HS diminished both cell surface binding and internalization of tau. In a cell culture, the addition of a 3-O-S HS 12-mer reduced both tau cell surface binding and cellular uptake. NMR titrations mapped 3-O-S binding sites to the microtubule binding repeat 2 (R2) and proline-rich region 2 (PRR2) of tau. Tau is only the seventh protein currently known to recognize HS 3-O-sulfation. Our work demonstrates that this rare 3-O-sulfation enhances tau–HS binding and likely the transcellular spread of tau, providing a novel target for disease-modifying treatment of AD and other tauopathies.     

Theoretical investigation into coordination and selectivity of uranyl-unilateral benzotriazole salophens (X = O/S) for R/S-triadimefons

The design of new uranyl-ligands (uranyl-Ls) is of great significance for the separation and utilization of uranium. In this paper, the triazole group was introduced into uranyl-salophen (uranyl-S) to form new asymmetric uranyl-unilateral benzotriazole salophen (uranyl-UBS); we further replaced two oxygen atoms of uranyl-UBS with two sulfur atoms to generate uranyl-unilateral benzotriazole thio-salophen (uranyl-UBTS) as a new receptor. Then, we comprehensively explored coordination models of uranyl-UBS and uranyl-UBTS with R/S-triadimefons (R/S-TDFs) enantiomers as the guests using density functional theory calculations at the B3LYP//RECP/6-311G** level; we then investigated enantioselectivity recognition of the new receptors to the guests R/S-TDFs. The results indicated that the uranium atoms of the receptors uranyl-S, uranyl-UBS and uranyl-UBTS could coordinate with the carbonyl oxygens in guests R/S-TDFs to form complexes of guest-receptors R/S-TDFs-uranyl-Ls that exhibited two stable V-shaped structures with quite different properties. It was found that the coordination ability to the guests R/S-TDFs is uranyl-UBTS > uranyl-UBS > uranyl-S, while the enantioselectivity for the guests is uranyl-UBTS > uranyl-S > uranyl-UBS and, when the receptor is the same, R-TDF has stronger coordination ability than S-TDF. These results provide information and theoretical supports for the experiments of asymmetric uranyl-UBS with R/S-TDFs, and produce a reference for further exploring the coordination characteristics of asymmetric uranyl-salophen with the triazole derivatives.     

The catalytic performance study of polymerized ionic liquid synthesized in different conditions on alkylation of o‐Xylene with styrene

In this work, a series of novel acidic polymerized ionic liquids were used as heterogeneous catalyst for alkylation of o‐Xylene with styrene. And the effect of the amount of initiator and the type of acid used for ion exchange on catalyst structure and the catalytic performance of catalysts for alkylation were studied thoroughly. The experiment results show: when the percentage of the amount of initiator in the total material is 3%, the polymerized ionic liquid catalyst MPM‐SO₃H‐[C₃V][SO₃CF₃] has the most uniform with a specific surface area of 97.30 m²/g and a pore volume of 0.35 cm³/g. Benefiting from the unique structure features, MPM‐SO₃H‐[C₃V][SO₃CF₃] manifested an excellent catalytic performance for alkylation of o‐Xylene with styrene, along with the conversion of styrene was 96.8% and the yield of 1‐Phenyl‐1‐ortho‐xylene ethane was 94.7%. Therefore, this work provides a novel reference to the synthesis of polymerized ionic liquids and clearly explains the advantage of novel acidic polymerized ionic liquids on alkylation.     

Click and count: specific detection of acid ceramidase activity in live cells

The use of intact cells in medical research offers a number of advantages over employing cell-free systems. In diagnostics, cells isolated from liquid biopsies can be directly used, speeding up the time of analysis and diminishing the risk of protein degradation by sample manipulation. In drug discovery, studies in live cells take into account aspects neglected in cell-free systems, such as uptake, metabolization, and subcellular concentration by compartmentalization of potential drug candidates. Therefore, probes for studies in cellulo are of paramount importance. Acid ceramidase (AC) is a lysosomal enzyme that hydrolyses ceramides into sphingoid bases and fatty acids. The essential role of this enzyme in the outburst and progress of several diseases, some of them still incurable, is well sustained. Despite the great clinical relevance of AC as a biomarker and therapeutic target, the specific monitoring of AC activity in live cells has remained elusive due to the concomitant existence of neutral and alkaline ceramidases. In this work, we report that 1-deoxydihydroceramides are exclusively hydrolysed by AC. Using N-octanoyl-18-azidodeoxysphinganine as a probe and a BODIPY-substituted bicyclononyne, we show the click-reliant predominant staining of lysosomes, with extra-lysosomal labeling also occurring in some cells. Importantly, using pharmacological and genetic tools together with high resolution mass spectrometry, we demonstrate that both lysosomal and extra-lysosomal staining are AC-dependent. These findings are translated into the specific flow cytometry monitoring of AC activity in intact cells, which fills an important gap in the field of diseases linked to altered AC activity.

The use of intact cells in medical research offers a number of advantages over employing cell-free systems.     

1,2-Diketones as photoinitiators of both cationic and free-radical photopolymerization under UV (392 nm) or Blue (455 nm) LEDs

The photoinitiation abilities of three 1,2-diketones [i.e., acenaphthenequinone ( ANPQ ), aceanthrenequinone ( AATQ ), and 9,10-phenanthrenequinone ( PANQ )]-based photoinitiating systems [PISs, with additives such as iodonium salt, N-vinylcarbazole (NVK), tertiary amine, and phenacyl bromide (R-Br)] for cationic photopolymerization and free-radical photopolymerization under the irradiation of ultraviolet (UV; 392 nm) or blue (455 nm) light-emitting diode (LED) bulb are investigated. All 1,2-diketones studied exhibit ground state absorption that match with the emission spectra of UV (392 nm) or blue LED (455 nm) better than that of the well-known blue-light-sensitive photoinitiator camphorquinone (CQ). In particular, AATQ /iodonium salt/NVK can show high photoinitiating ability (with epoxide conversion yield >70%) under the UV light irradiation due to the effect of NVK. In addition, 1,2-diketone/iodonium salt (and optional NVK) systems are capable of initiating free-radical photopolymerization of methacrylates, with conversions of 50–58%. Furthermore, some 1,2-diketone/tertiary amine (and optional R-Br) combinations are found to demonstrate high efficiency to initiate free-radical photopolymerization, and 71% of methacrylate conversion can be achieved with PANQ /tertiary amine/R-Br PIS. Some 1,2-ketone-based PISs can even exhibit higher efficiency than the CQ-based systems. The photochemical mechanism of the radical generation from the 1,2-diketone-based PISs is investigated and found to be consistent with the related photopolymerization efficiency. © 2020 Wiley Periodicals, Inc. J. Polym. Sci. 2020 , 58, 792–802     

Copper-Mediated Conversion of Complex Ethers to Esters: Enabling Biopolymer Depolymerisation under Mild Conditions

Selective processing of the β-O-4 unit in lignin is essential for the efficient depolymerisation of this biopolymer and therefore its successful integration into a biorefinery set-up. An approach is described in which this unit is modified to incorporate a carboxylic ester with the goal of enabling the use of mild depolymerisation conditions. Inspired by preliminary results using a Cu/TEMPO/O₂ system, a protocol was developed that gave the desired β-O-4-containing ester in high yield using certain dimeric model compounds. The optimised reaction conditions were then applied to an oligomeric lignin model system. Extensive 2D NMR analysis demonstrated that analogous chemistry could be achieved with the oligomeric substrate. Mild depolymerisation of the ester-containing oligomer delivered the expected aryl acid monomer.     

Synthesis,characterization, and thermal properties of novel silicon 1,1,3,3-tetramethylguanidinate derivatives and use as single-source chemical vapor deposition precursors

A series of chemical vapor deposition (CVD) precursors have been synthesized by a single-step reaction of 1,1,3,3-tetramethylguanidine and a variety of silicon chlorides. The structures of the 1,1,3,3-tetramethylguanidinate-based compounds were verified by ¹H NMR, ¹³C NMR, XPS, EI-MS, and elemental analysis. The thermal stability, transport behavior, and vapor pressures of these compounds were evaluated by simultaneous thermal analyses (STA). These compounds are highly stable and those in liquid form are very volatile. Silicon carbonitride (SiCN) thin films were prepared by using bis (tetramethylguanidine)-dimethyl-silane as the precursor in helicon wave plasma chemical vapor deposition (HWP-CVD). The properties of the films were investigated by SEM, AFM, and XPS. The results showed that the films have good uniformities, low friction coefficient, and high hardness, enabling the films for fabrication of semiconductor devices.