High-performance near-infrared OLEDs maximized at 925 nm and 1022 nm through interfacial energy transfer.

Using a transfer printing technique, we imprint a layer of a designated near-infrared fluorescent dye BTP-eC9 onto a thin layer of Pt(II) complex, both of which are capable of self-assembly. Before integration, the Pt(II) complex layer gives intense deep-red phosphorescence maximized at ~740 nm, while the BTP-eC9 layer shows fluorescence at > 900 nm. Organic light emitting diodes fabricated under the imprinted bilayer architecture harvest most of Pt(II) complex phosphorescence, which undergoes triplet-to-singlet energy transfer to the BTP-eC9 dye, resulting in high-intensity hyperfluorescence at > 900 nm. As a result, devices achieve 925 nm emission with external quantum efficiencies of 2.24% (1.94 ± 0.18%) and maximum radiance of 39.97 W sr-1 m-2. Comprehensive morphology, spectroscopy and device analyses support the mechanism of interfacial energy transfer, which also is proved successful for BTPV-eC9 dye (1022 nm), making bright and far-reaching the prospective of hyperfluorescent OLEDs in the near-infrared region.

Recognizing the Importance of Fast Nonisothermal Crystallization for High-Performance Two-Dimensional Dion-Jacobson Perovskite Solar Cells with High Fill Factors: A Comprehensive Mechanistic Study.

Two-dimensional (2D) Dion-Jacobson (DJ) perovskite solar cells (PSCs), despite their advantage in versatility of n-layer variation, are subject to poor photovoltaic efficiency, particularly in the fill factor (FF), compared to their three-dimensional counterparts. To enhance the performance of DJ PSCs, the process of growing crystals and hence the corresponding morphology of DJ perovskites are of prime importance. Herein, we report the fast nonisothermal (NIT) crystallization protocol that is previously unrecognized for 2D perovskites to significantly improve the morphology, orientation, and charge transport of the DJ perovskite films. Comprehensive mechanistic studies reveal that the NIT effect leads to the secondary crystallization stage, forming network-like channels that play a vital role in the FF's leap-forward improvement and hence the DJ PSC's performance. As a whole, the NIT crystallized PSCs demonstrate a high power conversion efficiency and an FF of up to 19.87 and 86.16%, respectively. This research thus provides new perspectives to achieve highly efficient DJ PSCs.

Cyano Derivatives of 7-Aminoquinoline That Are Highly Emissive in Water: Potential for Sensing Applications.

6-Cyano-7-aminoquinoline (6CN-7AQ) and 3-cyano-7-aminoquinoline (3CN-7AQ) were synthesized and found to exhibit intense emission with quantum yield as high as 63 % and 85 %, respectively, in water. Conversely, their derivatives 6-cyano-7-azidoquinoline (6CN-7N3 Q) and 3-cyano-7-azidoquinoline (3CN-7N3 Q) show virtually no emission, which makes them suitable to be used as recognition agents in azide reactions based on fluorescence recovery. Moreover, conjugation of 6CN-7AQ with a hydrophobic biomembrane-penetration peptide PFVYLI renders a nearly non-emissive 6CN-7AQ-PFVYLI composite, which can be digested by proteinase K, recovering the highly emissive 6CN-7AQ with ∼200-fold enhancement. The result provides an effective early confirmation for RT-qPCR in viral detection.

Unveiling the structural features of nonnative trimers of human superoxide dismutase 1.

BACKGROUND: Human SOD1 contains a single tryptophan residue (W32) which has been identified as a site of oxidative modification and a potentiator of aggregation involving in familial amyotrophic lateral sclerosis (fALS). In situ substitution of a tryptophan analog, 2,6-diazatryptophan ((2,6-aza)Trp) with its unique water-catalyzed proton transfer property, into proteins exhibits extraordinary sensitivity in the detection of subtle water-associated structural changes with only a few micro-molar concentration of samples. METHODS: A combination of size-exclusion chromatography and water-catalyzed fluorescent emission was utilized to probe the structural features of metastable SOD1 nonnative trimers, the potential neurotoxic species in the fALS. RESULTS: The monomer of apo-A4V SOD1 exhibits variable conformations and the fastest trimeric formation rate compared to that of wild type and I113T. The trimeric A4V SOD1 exhibits the least water molecules surrounding the W32, while I113T and the wild type appear to have more water molecules in the proximity of W32. A small molecule stabilizer, 5-fluorouridine, effects the structural conformation of SOD1 nonnative trimers. CONCLUSIONS: Our studies unveil new insights into water-associated structural changes of SOD1 nonnative trimers and demonstrate that in situ incorporation of (2,6-aza)Trp is a sensitive and powerful tool for probing subtle changes of water environments during protein aggregation. GENERAL SIGNIFICANCE: The water-sensitive probe, (2,6-aza)Trp, demonstrates superior sensitivity for detecting modulation of water microsolvation, structural conformation during oligomer formation and 5FUrd binding to both wild type and mutant SOD1.

The azatryptophan-based fluorescent platform for in vitro rapid screening of inhibitors disrupting IKKß-NEMO interaction.

The nuclear factor-κB (NF-κB) plays an important role in inflammatory and immune responses. Aberrant NF-κB signaling is implicated in multiple disorders, including cancer. Targeting the regulatory scaffold subunit IκB kinase γ (IKKγ/NEMO) as therapeutic interventions could be promising due to its specific involvement in canonical NF-κB activation without interfering with non-canonical signaling. In this study, the use of unnatural amino acid substituted IKKß with unique photophysical activity to sense water environment changes upon interaction with NEMO provides a powerful in vitro screening platform that would greatly facilitate the identification of compounds having the potential to disrupt IKKß-NEMO interaction, and thus specifically modulate the canonical NF-κB pathway. We then utilized a competitive binding platform to screen the binding ability of a number of potential molecules being synthesized. Our results suggest that a lead compound (-)-PDC-099 is a potent agent with ascertained potency to disrupt IKKß-NEMO complex for modulating NF-κB canonical pathway.

Unveiling the water-associated conformational mobility in the active site of ascorbate peroxidase.

We carried out comprehensive spectroscopic studies of wild type and mutants of ascorbate peroxidase (APX) to gain understanding of the conformational mobility of the active site. In this approach, three unnatural tryptophans were applied to replace the distal tryptophan (W41) in an aim to probe polarity/water environment near the edge of the heme-containing active site. 7-azatryptophan ((7-aza)Trp) is sensitive to environment polarity, while 2,7-azatryptophan ((2,7-aza)Trp) and 2,6-diazatryptophan ((2,6-aza)Trp) undergo excited-state water-catalyzed double and triple proton transfer, respectively, and are sensitive to the water network. The combination of their absorption, emission bands and the associated relaxation dynamics of these fluorescence probes, together with the Soret-band difference absorption and resonance Raman spectroscopy, lead us to unveil the water associated conformational mobility in the active site of APX. The results are suggestive of the existence of equilibrium between two different environments surrounding W41 in APX, i.e., the water-rich and water-scant forms with distinct fluorescence relaxation. Our results thus demonstrate for the first time the power of integrating multiple sensors (7-aza)Trp, (2,7-aza)Trp and (2,6-aza)Trp in probing the water environment of a specifically targeted Trp in proteins.

Snapshotting the Excited-State Planarization of Chemically Locked N,N'-Disubstituted Dihydrodibenzo[a,c]phenazines.

For deeper understanding of the coupling of electronic processes with conformational motions, we exploit a tailored strategy to harness the excited-state planarization of N,N'-disubstituted dihydrodibenzo[a,c]phenazines by halting the structural evolution via a macrocyclization process. In this new approach, 9,14-diphenyl-9,14-dihydrodibenzo[a,c]phenazine (DPAC) is used as a prototype, in which the para sites of 9,14-diphenyl are systematically enclosed by a dialkoxybenzene-alkyl-ester or -ether linkage with different chain lengths, imposing various degrees of constraint to impede the structural deformation. Accordingly, a series of DPAC-n (n = 1-8) derivatives were synthesized, in which n correlates with the alkyl length, such that the strength of the spatial constraint decreases as n increases. The structures of DPAC-1, DPAC-3, DPAC-4, and DPAC-8 were identified by the X-ray crystal analysis. As a result, despite nearly identical absorption spectra (onset ∼400 nm) for DPAC-1-8, drastic chain-length dependent emission is observed, spanning from blue (n = 1, 2, ∼400 nm) and blue-green (n = 3-5, 500-550 nm) to green-orange (n = 6) and red (n = 7, 8, ∼610 nm) in various regular solvents. Comprehensive spectroscopic and dynamic studies, together with a computational approach, rationalized the associated excited-state structure responding to emission origin. Severing the linkage for DPAC-5 via lipase treatment releases the structural freedom and hence results in drastic changes of emission from blue-green (490 nm) to red (625 nm), showing the brightening prospect of these chemically locked DPAC-n in both fundamental studies and applications.

The In Situ Tryptophan Analogue Probes the Conformational Dynamics in Asparaginase Isozymes.

Dynamic water solvation is crucial to protein conformational reorganization and hence to protein structure and functionality. We report here the characterization of water dynamics on the L-asparaginase structural homology isozymes L-asparaginases I (AnsA) and II (AnsB), which are shown via fluorescence spectroscopy and dynamics in combination with molecular dynamics simulation to have distinct catalytic activity. By use of the tryptophan (Trp) analog probe 2,7-diaza-tryptophan ((2,7-aza)Trp), which exhibits unique water-catalyzed proton-transfer properties, AnsA and AnsB are shown to have drastically different local water environments surrounding the single Trp. In AnsA, (2,7-aza)Trp exhibits prominent green N(7)-H emission resulting from water-catalyzed excited-state proton transfer. In stark contrast, the N(7)-H emission is virtually absent in AnsB, which supports a water-accessible and a water-scant environment in the proximity of Trp for AnsA and AnsB, respectively. In addition, careful analysis of the emission spectra and corresponding relaxation dynamics, together with the results of molecular dynamics simulations, led us to propose two structural states associated with the rearrangement of the hydrogen-bond network in the vicinity of Trp for the two Ans. The water molecules revealed in the proximity of the Trp residue have semiquantitative correlation with the observed emission spectral variations of (2,7-aza)Trp between AnsA and AnsB. Titration of aspartate, a competitive inhibitor of Ans, revealed an increase in N(7)-H emission intensity in AnsA but no obvious spectral changes in AnsB. The changes in the emission profiles reflect the modulation of structural states by locally confined environment and trapped-water collective motions.

Probing the polarity and water environment at the protein-peptide binding interface using tryptophan analogues.

7-Azatryptophan and 2,7-diazatryptophan are sensitive to polarity changes and water content, respectively, and should be ideal for studying protein-protein and protein-peptide interactions. In this study, we replaced the tryptophan in peptide Baa (LKWKKLLKLLKKLLKLG-NH2) with 7-azatryptophan or 2,7-diazatryptophan, forming (7-aza)Trp-Baa and (2,7-aza)Trp-Baa, to study the calmodulin (CaM)-peptide interaction. Dramatic differences in the (7-aza)Trp-Baa and (2,7-aza)Trp-Baa fluorescence properties between free peptide in water and calmodulin-bound peptide were observed, showing a less polar and water scant environment at the binding interface of the peptide upon calmodulin binding. The affinity of the peptides for binding CaM followed the trend Baa (210±10 pM)<(7-aza)Trp-Baa (109±5 pM)<(2,7-aza)Trp-Baa (45±2 pM), showing moderate increase in binding affinity upon increasing the number of nitrogen atoms in the Trp analogue. The increased binding affinity may be due to the formation of more hydrogen bonds upon binding CaM for the Trp analogue with more nitrogen atoms. Importantly, the results demonstrate that (7-aza)Trp and (2,7-aza)Trp are excellent probes for exploring the environment at the interface of protein-peptide interactions.

Induction of p53-independent growth inhibition in lung carcinoma cell A549 by gypenosides.

The objectives of this study are to investigate antiproliferative effect and mechanisms of bioactive compounds from Gynostemma pentaphyllum (G. pentaphyllum) on lung carcinoma cell A549. Saponins, carotenoids and chlorophylls were extracted and fractionated by column chromatography, and were subjected to high-performance liquid chromatography-mass spectrometry analyses. The saponin fraction, which consisted mainly of gypenoside (Gyp) XXII and XXIII, rather than the carotenoid and chlorophyll ones, was effective in inhibiting A549 cell growth in a concentration- and a time-dependent manner as evaluated using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. The estimated half maximal inhibitory concentration (IC50 ) of Gyp on A549 cells was 30.6 µg/ml. Gyp was further demonstrated to induce an apparent arrest of the A549 cell cycle at both the S phase and the G2/M phase, accompanied by a concentration- and a time-dependent increase in the proportions of both the early and late apoptotic cells. Furthermore, Gyp down-regulated cellular expression of cyclin A and B as well as BCL-2, while up-regulated the expression of BAX, DNA degradation factor 35 KD, poly [ADP-ribose] polymerase 1, p53, p21 and caspase-3. Nevertheless, both the treatment of a p53 inhibitor, pifithrin-α, and the small hairpin RNA-mediated p53 knockdown in the A549 cells did not alter the growth inhibition effect induced by Gyp. As a result, the cell cycle arrest and apoptosis of A549 cells induced by Gyp would most likely proceed through p53-independent pathway(s).

Probing water environment of Trp59 in ribonuclease T1: insight of the structure-water network relationship.

In this study, we used the tryptophan analogue, (2,7-aza)Trp, which exhibits water catalyzed proton transfer isomerization among N(1)-H, N(7)-H, and N(2)-H isomers, to probe the water environment of tryptophan-59 (Trp59) near the connecting loop region of ribonuclease Tl (RNase T1) by replacing the tryptophan with (2,7-aza)Trp. The resulting (2,7-aza)Trp59 triple emission bands and their associated relaxation dynamics, together with relevant data of 7-azatryptophan and molecular dynamics (MD) simulation, lead us to propose two Trp59 containing conformers in RNase T1, namely, the loop-close and loop-open forms. Water is rich in the loop-open form around the proximity of (2,7-aza)Trp59, which catalyzes (2,7-aza)Trp59 proton transfer in the excited state, giving both N(1)-H and N(7)-H isomer emissions. The existence of N(2)-H isomer in the loop-open form, supported by the MD simulation, is mainly due to the specific hydrogen bonding between N(2)-H proton and water molecule that bridges N(2)-H and the amide oxygen of Pro60, forming a strong network. The loop-close form is relatively tight in space, which squeezes water molecules out of the interface of α-helix and ß2 strand, joined by the connecting loop region; accordingly, the water-scant environment leads to the sole existence of the N(1)-H isomer emission. MD simulation also points out that the Trp-water pairs appear to preferentially participate in a hydrogen bond network incorporating polar amino acid moieties on the protein surface and bulk waters, providing the structural dynamic features of the connecting loop region in RNase T1.

Flavonoids from Gynostemma pentaphyllum exhibit differential induction of cell cycle arrest in H460 and A549 cancer cells.

Flavonoids, containing mainly kaempferol rhamnohexoside derivatives, were extracted from Gynostemma pentaphyllum (G. pentaphyllum) and their potential growth inhibition effects against H460 non-small cell lung cancer cells was explored and compared to that on A549 cells. The extracted flavonoids were found to exhibit antiproliferation effects against H460 cells (IC50 = 50.2 µg/mL), although the IC50 of H460 is 2.5-fold that of A549 cells (IC50 = 19.8 µg/mL). Further investigation revealed that H460 cells are more susceptible to kaempferol than A549, whereas A549 cell growth is better inhibited by kaempferol rhamnohexoside derivatives as compared with H460. In addition, flavonoids from G. pentaphyllum induced cell cycle arrest at both S and G2/M phases with concurrent modulated expression of the cellular proteins cyclin A, B, p53 and p21 in A549 cells, but not H460. On the contrary, apoptosis and concomitant alteration in balance of BCL-2 and BAX expression as well as activation of caspase-3 were equally affected between both cells by flavonoid treatment. These observations strongly suggest the growth inhibition discrepancy between H460 and A549 following flavonoid treatment can be attributed to the lack of cell cycle arrest in H460 cells and the differences between H460 and A549 cells may serve as contrasting models for further mechanistic investigations.

Probing water micro-solvation in proteins by water catalysed proton-transfer tautomerism.

Scientists have made tremendous efforts to gain understanding of the water molecules in proteins via indirect measurements such as molecular dynamic simulation and/or probing the polarity of the local environment. Here we present a tryptophan analogue that exhibits remarkable water catalysed proton-transfer properties. The resulting multiple emissions provide unique fingerprints that can be exploited for direct sensing of a site-specific water environment in a protein without disrupting its native structure. Replacing tryptophan with the newly developed tryptophan analogue we sense different water environments surrounding the five tryptophans in human thromboxane A2 synthase. This development may lead to future research to probe how water molecules affect the folding, structures and activities of proteins.

Probing ligand binding to thromboxane synthase.

Various fluorescence experiments and computer simulations were utilized to gain further understanding of thromboxane A(2) synthase (TXAS), which catalyzes an isomerization of prostaglandins H(2) to give rise to thromboxane A(2) along with a fragmentation reaction to 12-L-hydroxy-5,8,10-heptadecatrienoic acid and malondialdehyde. In this study, 2-p-toluidinylnaphthalene-6-sulfonic acid (TNS) was utilized as a probe to assess the spatial relationship and binding dynamics of ligand-TXAS interactions by steady-state and time-resolved fluorescence spectroscopy. The proximity between TNS and each of the five tryptophan (Trp) residues in TXAS was examined through the fluorescence quenching of Trp by TNS via an energy transfer process. The fluorescence quenching of Trp by TNS was abolished in the W65F mutant, indicating that Trp65 is the major contributor to account for energy transfer with TNS. Furthermore, both competitive binding experiments and the computer-simulated TXAS structure with clotrimazole as a heme ligand strongly suggest that TXAS has a large active site that can simultaneously accommodate TNS and clotrimazole without mutual interaction between TNS and heme. Displacement of TNS by Nile Red, a fluorescence dye sensitive to environmental polarity, indicates that the TNS binding site in TXAS is likely to be hydrophobic. The Phe cluster packing near the binding site of TNS may be involved in facilitating the binding of multiple ligands to the large active site of TXAS.

Construction and characterization of an electrospun tubular scaffold for small-diameter tissue-engineered vascular grafts: a scaffold membrane approach.

Based on a postulate that the microstructure of a scaffold can influence that of the resulting tissue and hence its mechanical behavior, we fabricated a small-diameter tubular scaffold (∼3 mm inner diameter) that has a microstructure similar to the arterial media using a scaffold membrane approach. Scaffold membranes that contain randomly oriented, moderately aligned, or highly aligned fibers were fabricated by collecting electrospun poly([epsilon]-caprolactone) fibers on a grounded rotating drum at three different drum rotation speeds (250, 1000, and 1500 rpm). Membranes of each type were wrapped around a small-diameter mandrel to form the tubular scaffolds. Particularly, the tubular scaffolds with three different off-axis fiber angles (30, 45, and 60 degree) were formed using membranes that contain aligned fibers. These scaffolds were subjected to biaxial mechanical testing to examine the effects of fiber directions as well as the distribution of fiber orientations on their mechanical properties. The circumferential elastic modulus of the tubular scaffold was closely related to the fiber directions; the larger the off-axis fiber angle the greater the circumferential elastic modulus. The distribution of fiber orientations, on the other hand, manifested itself in the mechanical behavior via the Poisson effect. Similar to cell sheet-based vascular tissue engineering, tubular cell-seeded constructs were prepared by wrapping cell-seeded scaffold membranes, alleviating the difficulty associated with cell seeding in electrospun scaffolds. Histology of the construct illustrated that cells were aligned to the fiber directions in the construct, demonstrating the potential to control the microstructure of tissue-engineered vascular grafts using the electrospun scaffold membrane.

Probing the interaction between prostacyclin synthase and prostaglandin H2 analogues or inhibitors via a combination of resonance Raman spectroscopy and molecular dynamics simulation approaches.

In an aim to probe the structure-function relationship of prostacyclin synthase (PGIS), resonance Raman (RR) spectroscopy and molecular dynamic (MD) simulation approaches have been exploited to characterize the heme conformation and heme-protein matrix interactions for human PGIS (hPGIS) and zebrafish PGIS (zPGIS) in the presence and absence of ligands. The high-frequency RR (1300-1700 cm(-1)) indicates that the heme group is in the ferric, six-coordinate, low-spin state for both resting and ligand-bound hPGIS/zPGIS. The low-frequency RR (300-500 cm(-1)) and MD simulation reveal a salient difference in propionate-protein matrix interactions between hPGIS and zPGIS, as evident by a predominant propionate bending vibration at 386 cm(-1) in resting hPGIS, but two vibrations near 370 and 387 cm(-1) in resting zPGIS. Upon binding of a substrate analogue (U46619, U51605, or U44069), both hPGIS and zPGIS induce a distinctive perturbation of the propionate-protein matrix interactions, resulting in similar Raman shifts to ~381 cm(-1). On the contrary, the bending vibration remains unchanged upon binding of inhibitor/ligand (minoxidil, clotrimazole, or miconazole), indicating that these inhibitors/ligands do not interfere with the propionate-protein matrix interactions. These results, together with subtle changes in vinyl bending modes, demonstrate drastically different RR shifts with heme conformational changes in both hPGIS and zPGIS upon different ligand bindings, suggesting that PGIS exhibits a ligand-specific heme conformational change to accommodate the substrate binding. This substrate-induced modulation of the heme conformation may confer high product fidelity upon PGIS catalysis.