Ring Subunits Research Articles

Simple synthesis of hierarchical Co-Meso-UTL zeolite for efficient epoxidation of styrene

Co-zeolites with hierarchical porous are significantly desirable for the liquid-phase selective epoxidation of styrene that required relatively open reaction spaces. A typical UTL germanosilicate with the absence of organic structure-directing agent molecules inside pore channels suffered from severe structural collapse after ultrasonic treatment under pure water medium, originating from the unstable germanium-rich double four ring subunits. The simple strategy cleaved the weak linkage Ge-O bands and caused the irregular location of silicon-rich cfi layers. Subsequently, Co activity centers was incorporated into the framework through reacting with the abundant silanol groups located in the hydroxyl nests at room temperature. The random arrangement of mono- or multi- cfi layers induced the stacking faults and generated a large amount of intracrystal mesopores after calcination. The resultant Co-Meso-UTL catalysts favored the molecules diffusion of large-sized styrene and target products, improving the accessibility of Co active sites and exhibiting excellent catalytic performance in the styrene epoxidation reaction with single hydrogen peroxide (H2O2) oxidant. Especially, high styrene oxide product selectivity was also obtained through controlling appropriate H2O2-added content as well as reaction time, which possessed a highest turnover frequency value of 403 h−1 among the reported catalysts.

Geometric Structure, Electronic, and Spectral Properties of Metal-free Phthalocyanine under the External Electric Fields.

Here, the ground-state structures, electronic structures, polarizability, and spectral properties of metal-free phthalocyanine (H2Pc) under different external electric fields (EEFs) are investigated. The results show that EEF has an ultrastrong regulation effect on various aspects of H2Pc; the geometric structures, electronic properties, polarizability, and spectral properties are strongly sensitive to the EEF. In particular, an EEF of 0.025 a.u. is an important control point: an EEF of 0.025 a.u. will bend the benzene ring subunits to the positive and negative x directions of the planar molecule. Flipping the EEF from positive (0.025 a.u.) to negative (-0.025 a.u.) flips also the bending direction of benzene ring subunits. The H2Pc shows different dipole moments projecting an opposite direction along the x direction (-84 and 84 Debye for EEFs of -0.025 and 0.025 a.u., respectively) under negative and positive EEF, revealing a significant dipole moment transformation. Furthermore, when the EEF is removed, the molecule can be restored to the planar structure. The transformation of the H2Pc structure can be induced by the EEF, which has potential applications in the molecular devices such as molecular switches or molecular forceps. EEF lowers total energy and reduces highest occupied molecular orbital-lowest unoccupied molecular orbital (HOMO-LUMO) gap; especially, an EEF of 0.025 a.u. can reduce the HOMO-LUMO gap from 2.1 eV (in the absence of EEF) to 0.37 eV, and thus, it can enhance the molecular conductivity. The first hyperpolarizability of H2Pc is 0 in the absence of EEF; remarkably, an EEF of 0.025 a.u. can enhance the first hyperpolarizability up to 15,578 a.u. Therefore, H2Pc under the EEF could be introduced as a promising innovative nonlinear optical (NLO) nanomaterial such as NLO switches. The strong EEF (0.025 a.u.) causes a large number of new absorption peaks in IR and Raman spectra and causes the redshift of electronic absorption spectra. The changes of EEF can be used to regulate the structure transformation and properties of H2Pc, which can promote the application of H2Pc in nanometer fields such as molecular devices.

Planar Si5 and Ge5 Pentagons beside Isolated Phosphide Anions in Lithium Phosphide Tetrelides Li10+xSi5P and Li10+xGe5P

AbstractBinary lithium silicides and germanides as well as ternary lithium phosphidotetrelates have been intensively investigated with regard to their suitability as anode materials for all‐solid‐state‐batteries. In this context we studied the ternary phase diagram Li/Tt/P on the phosphorus‐poor side. The phosphide tetrelides Li10.68(8)Si5P and Li10.1(2)Ge5P are ternary compounds in the respective phase systems so far uniquely featuring discrete building blocks, which were previously found exclusively in binary alkali metal tetrelides and phosphides. Li10.68(8)Si5P and Li10.1(2)Ge5P contain aromatic Tt5 pentagons with an environment of mostly ordered Li atoms and isolated P3− anions with a coordination environment of predominantly disordered Li atoms. The five‐membered ring subunits are stacked under formation of [Li6Tt5] chains (Tt=Si, Ge). chains. The compounds crystallize in the space group Pnma (63) (Z=4) with cell parameters of a=10.2434(3) Å, b=4.2788(1) Å, c=23.9767(3) Å for Li10.68(8)Si5P and a=10.3599(11) Å, b=4.3072(2) Å, c=24.267(2) Å for Li10.1(2)Ge5P. The single crystal structure determination is supplemented by X‐ray powder diffraction experiments and 7Li, 29Si and 31P solid state MAS NMR spectroscopy. With respect to their electron count the compounds are placed between compounds comprising aromatic five‐membered rings such as the electron‐precise Zintl phase Li8MgSi6 and the non‐classical Zintl phase Li12Si7 having two extra electrons.

Low tolerance for transcriptional variation at cohesin genes is accompanied by functional links to disease-relevant pathways

Background: The cohesin complex plays an essential role in genome organisation and cell division. A full complement of the cohesin complex and its regulators is important for normal development, since...

Dipodal, Tripodal, and Discoidal Coordination Modes of Kemp's Triacid Anions

In order to probe the conformational preferences of the anions of Kemp's triacid (cis,cis‐1,3,5‐trimethyl‐1,3,5‐cyclohexanetricarboxylic acid, H3kta) in their metal ion complexes, and their influence on the structures thereof, six complexes with widely different metal cations (alkali, alkaline‐earth, 3d‐block and actinide) were synthesized and crystallographically characterized. All these complexes crystallize as diperiodic assemblies, but with three different conformations of the kta3–/Hkta2–/H3kta ligands. Only the tripodal form, with the cyclohexyl ring in chair conformation and the three carboxylic/ate groups axial is found in [Na5(kta)(Hkta)(H2O)] (1), [Sr3(H3kta)(kta)2] (2), [Mn3(kta)2(H2O)3] (3), and [Mn(H2O)6][Mn5(kta)4(H2O)4] (4). Particularly notable are hexanuclear ring subunits formed around two ligands in 2. The tripodal form coexists with the discoidal one, with the ring in the chair conformation and the three carboxylate groups equatorial, in [Mn6(kta)4(nmp)3] (5), with nmp = N‐methyl‐2‐pyrrolidone, which displays hexanuclear ring subunits. Finally, the so‐called dipodal form, with the ring in boat conformation, two carboxylate groups axial and one equatorial, is only found in [UO2Cs(kta)] (6), in which uranyl cations and ligands alone form a honeycomb assembly, the caesium cations being decorating species.

Improved structural stability and catalytic performance for CH4 reforming reaction of mesoporous MCM-41 analogs prepared by zeolitic subunits of MOR desilication

We demonstrated a new strategy for the preparation of highly stable mesoporous molecular sieves (TMM) by the desilication of MOR zeolite and self-assembling of the zeolite subunits in Na2O•(3.3–3.4) SiO2 aqueous solution. Na2O•(3.3–3.4) SiO2 in the aqueous solution plays two roles in the synthesis, one was providing a moderate alkaline medium with buffering effect for the desilication of MOR zeolite, other one was supplementing silica source. The desilication conduction of MOR zeolite was optimized by the adjustment of reaction temperature and time, pH in the solution. TMM were characterized by X-ray diffraction (XRD), N2 adsorption/desorption (BET), Fourier transformed infrared (FT-IR) and transmission electron microscopy (TEM). The characterization results showed that the characteristics of MOR zeolite, such as double six-membered or five-membered ring subunits, were still preserved in TMM, and TMM possessed well-ordered mesostructure of MOR zeolite crystals. Compared with the pure silica mesoporous molecular sieve (MCM-41, 4-day), the hydrothermal stability of TMM (17-day) was increased by 100% with refluxed water. The significantly improving of TMM hydrothermal stability was due to increase the surface hydrophobicity and the ratio of T12-ring, and decreased remarkably number of hydrophilic silanol groups. After hydrothermal treatment for 17-day, TMM also exhibited more efficient catalytic activation than MCM-41 treated for 4-day in CO2 reforming of methane reaction (700 °C). The 5 wt% Ni/TMM-17d-Glu (5 wt% Ni, loading) catalysts prepared by Glucose modified impregnation method exhibited excellent catalytic activity, which were providing 58% CO2 and 47% CH4 conversions at 700 °C.

Mutations in multiple components of the nuclear pore complex cause nephrotic syndrome.

Steroid-resistant nephrotic syndrome (SRNS) almost invariably progresses to end-stage renal disease. Although more than 50 monogenic causes of SRNS have been described, a large proportion of SRNS remains unexplained. Recently, it was discovered that mutations of NUP93 and NUP205, encoding 2 proteins of the inner ring subunit of the nuclear pore complex (NPC), cause SRNS. Here, we describe mutations in genes encoding 4 components of the outer rings of the NPC, namely NUP107, NUP85, NUP133, and NUP160, in 13 families with SRNS. Using coimmunoprecipitation experiments, we showed that certain pathogenic alleles weakened the interaction between neighboring NPC subunits. We demonstrated that morpholino knockdown of nup107, nup85, or nup133 in Xenopus disrupted glomerulogenesis. Re-expression of WT mRNA, but not of mRNA reflecting mutations from SRNS patients, mitigated this phenotype. We furthermore found that CRISPR/Cas9 knockout of NUP107, NUP85, or NUP133 in podocytes activated Cdc42, an important effector of SRNS pathogenesis. CRISPR/Cas9 knockout of nup107 or nup85 in zebrafish caused developmental anomalies and early lethality. In contrast, an in-frame mutation of nup107 did not affect survival, thus mimicking the allelic effects seen in humans. In conclusion, we discovered here that mutations in 4 genes encoding components of the outer ring subunits of the NPC cause SRNS and thereby provide further evidence that specific hypomorphic mutations in these essential genes cause a distinct, organ-specific phenotype.

Cohesin SA2 is a sequence-independent DNA-binding protein that recognizes DNA replication and repair intermediates

Proper chromosome alignment and segregation during mitosis depend on cohesion between sister chromatids, mediated by the cohesin protein complex, which also plays crucial roles in diverse genome maintenance pathways. Current models attribute DNA binding by cohesin to entrapment of dsDNA by the cohesin ring subunits (SMC1, SMC3, and RAD21 in humans). However, the biophysical properties and activities of the fourth core cohesin subunit SA2 (STAG2) are largely unknown. Here, using single-molecule atomic force and fluorescence microscopy imaging as well as fluorescence anisotropy measurements, we established that SA2 binds to both dsDNA and ssDNA, albeit with a higher binding affinity for ssDNA. We observed that SA2 can switch between the 1D diffusing (search) mode on dsDNA and stable binding (recognition) mode at ssDNA gaps. Although SA2 does not specifically bind to centromeric or telomeric sequences, it does recognize DNA structures often associated with DNA replication and double-strand break repair, such as a double-stranded end, single-stranded overhang, flap, fork, and ssDNA gap. SA2 loss leads to a defect in homologous recombination-mediated DNA double-strand break repair. These results suggest that SA2 functions at intermediate DNA structures during DNA transactions in genome maintenance pathways. These findings have important implications for understanding the function of cohesin in these pathways.

Solvent impact on the planarity and aromaticity of free and monohydrated zinc phthalocyanine: a theoretical study

A theoretical investigation on the planarity of molecular structure of zinc phthalocyanine (ZnPc) and its aromaticity has been performed using B3LYP and M06-2X density functionals combined with selected Pople-type basis sets. The effect of the applied calculation method on the optimized structure of ZnPc and ZnPc∙∙∙H2O, both in the gas phase and in the polar solvent, was analyzed. To quantify the aromaticity of the ZnPc and ZnPc∙∙∙H2O complexes, both the geometric and magnetic criteria, i.e., Harmonic Oscillator Model of Aromaticity (HOMA) index and the nucleus-independent chemical shift (NICS) values at the centers or 1 Å above the centers of structural subunits, were calculated. The energies of highest energy occupied molecular orbital (HOMO) and lowest energy unoccupied molecular orbital (LUMO) and energy gaps were also estimated. The results show that the free ZnPC molecule is flat in the gas phase and nonplanar in the polar environments (DMSO and water). ZnPC∙∙∙H2O is nonpolar in the gas phase and polar solvent which is in agreement with recent X-ray reports. Both HOMA and NICS indexes indicate the presence of highly aromatic macrocycle and benzene rings while these parameters for pyrrolic ring are significantly smaller than in free pyrrole. The presence of polar solvents practically does not change aromaticity of the ring subunits of the studied compounds.

Aerosol Cross-Linked Crown Ether Diols Melded with Poly(vinyl alcohol) as Specialized Microfibrous Li+ Adsorbents.

Crown ether (CE)-based Li+ adsorbent microfibers (MFs) were successfully fabricated through a combined use of CE diols, electrospinning, and aerosol cross-linking. The 14- to 16-membered CEs, with varied ring subunits and cavity dimensions, have two hydroxyl groups for covalent attachments to poly(vinyl alcohol) (PVA) as the chosen matrix. The CE diols were blended with PVA and transformed into microfibers via electrospinning, a highly effective technique in minimizing CE loss during MF fabrication. Subsequent aerosol glutaraldehyde (GA) cross-linking of the electrospun CE/PVA MFs stabilized the adsorbents in water. The aerosol technique is highly effective in cross-linking the MFs at short time (5 h) with minimal volume requirement of GA solution (2.4 mL g-1 MF). GA cross-linking alleviated CE leakage from the fibers as the CEs were securely attached with PVA through covalent CE-GA-PVA linkages. Three types of CE/PVA MFs were fabricated and characterized through Fourier transform infrared-attenuated total reflection, 13C cross-polarization magic angle spinning NMR, field emission scanning electron microscope, N2 adsorption/desorption, and universal testing machine. The MFs exhibited pseudo-second-order rate and Langmuir-type Li+ adsorption. At their saturated states, the MFs were able to use 90-99% CEs for 1:1 Li+ complexation, suggesting favorability of their microfibrous structures for CE accessibility to Li+. The MFs were highly Li+-selective in seawater. Neopentyl-bearing CE was most effective in blocking larger monovalents Na+ and K+, whereas the dibenzo CE was best in discriminating divalents Mg2+ and Ca2+. Experimental selectivity trends concur with the reaction enthalpies from density functional theory calculations, confirming the influence of CE structures and cavity dimensions in their "size-match" Li+ selectivity.

The elastic transfer model of angular rate modulation in F1-ATPase stalling and controlled rotation experiments

The recent experimental, theoretical and computational advances in the field of F1-ATPase single-molecule microscopy are briefly surveyed. The role of theory is revealed in the statistical analysis, interpretation and prediction of single-molecule experimental trajectories, and in linking them with atomistic simulations. In particular, a theoretical model of elastically coupled molecular group transfer is reviewed and a detailed method for its application in stalling and controlled rotation experiments is provided. It is shown how the model can predict, using previous experiments, the rates of ligand binding/release processes (steps) and their exponential dependence on rotor angle in these experiments. The concept of Brønsted slopes is reviewed in the context of the single-molecule experiments, and the rate versus rotor angle relations are explained using the elastic model. These experimental data are treated in terms of the effect of thermodynamic driving forces on the rates assuming that the rotor shaft is elastically coupled to stator ring subunits in which the steps occur. In the application of the group transfer model on an extended angular range processes leading up to the transfer are discussed. Implications for large-scale atomistic simulation are suggested for the treatment of torque-generating steps.

Ab initio computations of zero-field splitting parameters and effective exchange integrals for single-molecule magnets (Mn12- and Mn11Cr-acetate clusters)

In this study, the zero-field splitting parameters (D, E) and effective exchange integrals (J) for a Mn12/Mn11Cr mixed crystal were evaluated using ab initio molecular orbital (MO) computations based on the hybrid density functional theory (DFT) methods for Mn11Cr∗-acetate cluster (Mn12 and Mn11Cr mixing). In this study, Mn12-acetate, Mn11Cr-acetate, and Mn12−nCrn-acetate clusters were focused on. The zero-field splitting (ZFS) parameters are obtained by UB3LYP methods, i.e. −0.339cm−1 (Mn12-acetate) and −0.351cm−1 (Mn11Cr-acetate). The energy barriers were estimated to be 53.7K and 50.4K, which showed good agreement with previously reported experimental values of 68.8K and 56.8K, respectively. The origin of such anisotropy is assumed to be the eight MnIII ions with d4-electrons in their outer ring subunits, where elongated conformation along the axial direction becomes very important and leads to negative D values. The spin networks in these clusters were discussed by using the effective exchange integral sets. The Js value, as estimated by the UB3LYP method revealed that the J2, J3, and J3′ interactions played important roles in describing the parallel spin ordering in cubane and ring subunits as well as the antiferromagnetic coupling between two subunits. The other interactions, i.e. J1, J1′, J4, and J4′ somewhat disturbed the spin ordering in the Mn12-acetate cluster. The substitution of MnIV by CrIII did not change the spin network. However, the total S value changed after the substitution.

Mesopore modification of beta zeolites by sequential alkali and acid treatments: Narrowing mesopore size distribution featuring unimodality and mesoporous texture properties estimated upon a mesoporous volumetric model

For mesopore modification of zeolite beta prepared in protonic form (H-Beta, Si/Al = ca.18.7) to concentrate pore size distributions, the framework T-atoms of H-Beta crystals were extracted according to the sequential schemes: hydrothermal pre-etching using incipiently wetted alkaline solutions, bulk alkaline treatment, and acid leaching coupling with ammonium exchange. Based on XRD, FTIR, N2 adsorption at 77 K, SEM, TEM, EDX, ICP–AES, and immersion porosimetry, structure properties of the hierarchical materials obtained were correlated with the sequential treatment processes. In contrast to single bulk alkaline treatment, the hydrothermal alkali pre-etching enables to impair or eliminate broadly secondary pore–distributed peaks, the bulk alkaline treatment before acid leaching facilitates textural mesopore formation, and the final acid leaching plus ammonium exchange leads to moderate increases in mesoporosity and mesopore diameters, and also to a suitable enhancement of Si/Al ratios. FTIR spectra provided evidence for the short range ordering of modified zeolites and for the presence of zeolitic ring sub-units in the basic filtrate. However, the microporosity, crystallinity and yield relative to the parent are reduced by increased alkaline concentrations. A novel model was built for estimating a volumetric fraction of intracrystalline-to N2 adsorbed mesopores. Calculation results show that this volumetric fractional model promises to identify the mesoporous textural property, explaining how many volumetric fractions of mesopores are to locate in beta crystallite entities. In addition, the hierarchical beta possesses a mesopore network with higher hydrothermal stability at 873 K.

Palladium-catalysed cyclisation of alkenols: Synthesis of oxaheterocycles as core intermediates of natural compounds.

The study of Pd-catalysed cyclisation reactions of alkenols using different catalytic systems is reported. These transformations affect the stereoselective construction of mono- and/or bicyclic oxaheterocyclic derivatives depending on a starting alkenol. The substrate scope and proposed mechanism of Pd-catalysed cyclisation reactions are also discussed. Moreover, the diastereoselective Pd-catalysed cyclisation of appropriate alkenols to tetrahydrofurans and subsequent cyclisation provided properly substituted 2,5-dioxabicyclo[2.2.1]heptane and 2,6-dioxabicyclo[3.2.1]octane, respectively. Such bicyclic ring subunits are found in many natural products including ocellenynes and aurovertines.

Construction of several new s-/p-block complexes containing binuclear metal–terpyridine building blocks: dependence of structural diversity on the number of coordinated water molecules

Five new coordination polymers, [Mg(DSPT)(H2O)2]·2H2O (H2DSPT = 4′-(2,4-disulfophenyl)-2,2′:6′2′′-terpyridine) (1), {[Ca(DSPT)(H2O)2]·1.5H2O}n (2), {[Pb(DSPT)(H2O)]·2H2O}n (3), {[Pb(DSPT)]·0.5H2O}n (4), and {[Pb3(DSPT)3(H2O)]·8H2O}n (5), were prepared under hydrothermal conditions. They are all complexes containing similar M2L2 binuclear metal–terpyridine building units. Complex 1 is a discrete binuclear molecule that extends to 2D layers via O–H⋯O hydrogen bonds between coordinated water and a sulfo group on H2DSPT. Complexes 2 and 3 are 1D coordination chains based on [M2(DSPT)2] ring subunits but are connected by different M–O bonds, and the 1D chains are also further connected by O–H⋯O hydrogen bonds between coordinated water and a sulfo group on H2DSPT to a 2D layer. Complex 4 exhibits a 2D coordination network also based on the [M2(DSPT)2] ring subunits. Complex 5 is a by-product of the synthesis of complex 4 that displays a 1D coordination chain structure containing the same [M2(DSPT)2] ring subunits. Interestingly, the 2D layers described above show identical skeletons when the bond types between [M2(DSPT)2] ring subunits are neglected. The 2D layers in complexes 1–3 can be derived from replacing selected coordination bonds in complex 4 with hydrogen bonds, and such structural diversity is largely dependent on the number of coordinated water molecules around the metal center. Thermogravimetric analyses of 1–4 showed that they were stable at temperatures higher than 500 °C. The luminescent properties of complexes 1–4 were also investigated.

SA1 binds directly to DNA via its unique AT-hook to promote sister chromatid cohesion at telomeres

Sister chromatid cohesion relies on cohesin, a complex comprising a tri-partite ring and a peripheral subunit Scc3, which is found as two related isoforms SA1 and SA2 in vertebrates. There is a division of labor between the vertebrate cohesin complexes; SA1-cohesin is required at telomeres and SA2-cohesin at centromeres. Depletion of SA1 has dramatic consequences for telomere function and genome integrity, but the mechanism by which SA1-cohesin mediates cohesion at telomeres is not well understood. Here we dissect the individual contribution of SA1 and the ring subunits to telomere cohesion and show that telomeres rely heavily on SA1 and to a lesser extent on the ring for cohesion. Using chromatin immunoprecipitation we show that SA1 is highly enriched at telomeres, is decreased at mitosis when cohesion is resolved, and is increased when cohesion persists. Overexpression of SA1 alone was sufficient to induce cohesion at telomeres, independent of the cohesin ring and dependent on its unique (not found in SA2) N-terminal domain, which we show binds to telomeric DNA through an AT-hook motif. We suggest that a specialized cohesion mechanism may be required to accommodate the high level of DNA replication-associated repair at telomeres.

High Degree of Coordination and Division of Labor among Subunits in a Homomeric Ring ATPase

Ring NTPases of the ASCE superfamily perform a variety of cellular functions. An important question about the operation of these molecular machines is how the ring subunits coordinate their chemical and mechanical transitions. Here, we present a comprehensive mechanochemical characterization of a homomeric ring ATPase-the bacteriophage φ29 packaging motor-a homopentamer that translocates double-stranded DNA in cycles composed of alternating dwells and bursts. We use high-resolution optical tweezers to determine the effect of nucleotide analogs on the cycle. We find that ATP hydrolysis occurs sequentially during the burst and that ADP release is interlaced with ATP binding during the dwell, revealing a high degree of coordination among ring subunits. Moreover, we show that the motor displays an unexpected division of labor: although all subunits of the homopentamer bind and hydrolyze ATP during each cycle, only four participate in translocation, whereas the remaining subunit plays an ATP-dependent regulatory role.

Characterization and investigation mechanism of hydrothermal stability for micropore-free ordered mesoporous molecular sieves

SBA-15-type mesoporous aluminosilicates molecular sieves (MSAMS-2 and MSAMS-4) were synthesized and characterized by N2 adsorption–desorption, XRD, FT-IR, TG/DTA, FE-SEM, TEM and HR-TEM. N2 adsorption–desorption and t-plots of N2 adsorptions show the two sieves are free of micropores within their mesoporous framework wall. MSAMS-2 and MSAMS-4 exhibit extraordinary stability both in boiling water (over 360h) and in high temperature steam (800°C for 6h). FT-IR together with XRD, TG/DTA, HR-TEM and FE-SEM characterizations demonstrate that five-membered ring subunits with small size rather than zeolite crystals were only formed within the framework pores of MSAMS-2 and MSAMS-4. These five-membered ring subunits are extraordinarily stable and can effectively keep them from water molecule attack, which should play a crucial role in improving the hydrothermal stability for MSAMS-2 and MSAMS-4.

Engineered Vascular Tissue Fabricated from Aggregated Smooth Muscle Cells

The goal of this study was to develop a system to rapidly generate engineered tissue constructs from aggregated cells and cell-derived extracellular matrix (ECM) to enable evaluation of cell-derived tissue structure and function. Rat aortic smooth muscle cells seeded into annular agarose wells (2, 4 or 6 mm inside diameter) aggregated and formed thick tissue rings within 2 weeks of static culture (0.76 mm at 8 days; 0.94 mm at 14 days). Overall, cells appeared healthy and surrounded by ECM comprised of glycosoaminoglycans and collagen, although signs of necrosis were observed near the centers of the thickest rings. Tissue ring strength and stiffness values were superior to those reported for engineered tissue constructs cultured for comparable times. The strength (100–500 kPa) and modulus (0.5–2 MPa) of tissue rings increased with ring size and decreased with culture duration. Finally, tissue rings cultured for 7 days on silicone mandrels fused to form tubular constructs. Ring margins were visible after 7 days, but tubes were cohesive and mechanically stable, and histological examination confirmed fusion between ring subunits. This unique system provides a versatile new tool for optimization and functional assessment of cell-derived tissue, and a new approach to creating tissue-engineered vascular grafts.

The Steady-State Repertoire of Human SCF Ubiquitin Ligase Complexes Does Not Require Ongoing Nedd8 Conjugation

The human genome encodes 69 different F-box proteins (FBPs), each of which can potentially assemble with Skp1-Cul1-RING to serve as the substrate specificity subunit of an SCF ubiquitin ligase complex. SCF activity is switched on by conjugation of the ubiquitin-like protein Nedd8 to Cul1. Cycles of Nedd8 conjugation and deconjugation acting in conjunction with the Cul1-sequestering factor Cand1 are thought to control dynamic cycles of SCF assembly and disassembly, which would enable a dynamic equilibrium between the Cul1-RING catalytic core of SCF and the cellular repertoire of FBPs. To test this hypothesis, we determined the cellular composition of SCF complexes and evaluated the impact of Nedd8 conjugation on this steady-state. At least 42 FBPs assembled with Cul1 in HEK 293 cells, and the levels of Cul1-bound FBPs varied by over two orders of magnitude. Unexpectedly, quantitative mass spectrometry revealed that blockade of Nedd8 conjugation led to a modest increase, rather than a decrease, in the overall level of most SCF complexes. We suggest that multiple mechanisms including FBP dissociation and turnover cooperate to maintain the cellular pool of SCF ubiquitin ligases.