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A new molecular beacon (MB) driven by two-photon excitation (TPE) using quantum dots as energy donor is constructed, which provides reduced direct excitation of acceptor and is free of interferences from autofluorescence or scattering light in a complicated biological matrix. [ABSTRACT FROM AUTHOR]

This study features solid phase syntheses of cyclic tetrapeptides containing anthranilic acid (Anth) on relatively inexpensive resins derived from polystyrene. It proved to be difficult to hydrolyze a supported Anth-methyl ester unless a phase transfer catalyst was added to facilitate transport of hydroxide into the swollen hydrophobic gel state of the resin. We suggest this may be an under-appreciated strategy for improving syntheses on polystyrene supports. [ABSTRACT FROM AUTHOR]

Abstract Active yet low‐cost electrocatalysts for water oxidation are crucial for the development of hydrogen energy economy. The Fe doping into Ni(OH)2 dramatically enhances catalytic activity toward oxygen evolution reaction (OER) but fabricating Ni(OH)2 of high Fe loading is still challenging. Herein, we report a one‐pot strategy to prepare disordered β‐Ni(OH)2 nanosheets with a high Fe doping level (9.9 at%, D‐Fe‐Ni(OH)2). By engaging 1,4‐phenylenediphosphonic acid (BDPA), FexBDPAy precursors are in situ generated in a growth solution containing Fe3+ ions, which decrease the reaction kinetics of Ni2+ and Fe3+ ions at the surface of Ni foam. This prevents the deconstructive hydrolysis by Fe3+ ions and enables a high Fe‐doping in D‐Fe‐Ni(OH)2. The as‐prepared D‐Fe‐Ni(OH)2 affords 10 mA cm−2 at an ultralow OER overpotential of 194 mV in alkaline media. This work offers a promising strategy of engaging organic ligands to achieve high‐doping levels for the construction of efficient electrocatalysts.

FtsQBL is a transmembrane protein complex in the divisome of Escherichia coli that plays a critical role in regulating cell division. Although extensive efforts have been made to investigate the interactions between the three involved proteins, FtsQ, FtsB, and FtsL, the detailed interaction mechanism is still poorly understood. In this study, we used hydrogen-deuterium exchange mass spectrometry to investigate these full-length proteins and their complexes. We also dissected the structural dynamic changes and the related binding interfaces within the complexes. Our data revealed that FtsB and FtsL interact at both the periplasmic and transmembrane regions to form a stable complex. Furthermore, the periplasmic region of FtsB underwent significant conformational changes. With the help of computational modeling, our results suggest that FtsBL complexation may bring the respective constriction control domains (CCDs) in close proximity. We show that when FtsBL adopts a coiled-coil structure, the CCDs are fixed at a vertical position relative to the membrane surface; thus, this conformational change may be essential for FtsBL's interaction with other divisome proteins. In the FtsQBL complex, intriguingly, we show only FtsB interacts with FtsQ at its C-terminal region, which stiffens a large area of the β-domain of FtsQ. Consistent with this, we found the connection between the α- and β-domains in FtsQ is also strengthened in the complex. Overall, the present study provides important experimental evidence detailing the local interactions between the full-length FtsB, FtsL, and FtsQ protein, as well as valuable insights into the roles of FtsQBL complexation in regulating divisome activity. [ABSTRACT FROM AUTHOR]

The imaging capability of conventional lenses is mainly limited by the diffraction of light, and the so-called superlens has been developed allowing the recovery of evanescent waves in the focal plane. However, the remarkable focusing behavior of the superlens is greatly confined in the near-field regime due to the exponential decay of evanescent waves. To tackle this issue, we design a waveguide metasurface-based superlens with an extraordinary quasi-far-field focusing capability beyond the diffraction limit in the present work. Specifically, we analyze the underlying physical mechanism and provide experimental verification of the proposed superlens. The metasurface superlens is formed by an array of gradient nanoslits perforated in a gold slab, and supports transverse-electric (TE) waveguide modes under linearly polarized illumination along the long axis of the slits. Numerical results illustrate that exciting such TE waveguide modes can modulate not only optical phase but also evanescent waves. Consequently, some high-spatial-frequency waves can contribute to the focusing of the superlens, leading to the quasi-far-field super-resolution focusing of light. Under 405 nm illumination and oil immersion, the fabricated superlens shows a focus spot of 98 nm (i.e. λ/4.13) at a focal distance of 1.49 μm (i.e. 3.68λ) using an oil immersion objective, breaking the diffraction limit of λ/2.38 in the quasi-far field regime. The developed metasurface optical superlens with such extraordinary capabilities promises exciting avenues to nanolithography and ultra-small optoelectronic devices.

Chemical fuels, produced from light, afford an alternative to fossil fuel, but conversion materials suffer from low photon-to-fuel efficiencies. Here, authors incorporate gold/N-doped TiO2 on MgO surfaces and show enhanced photocatalytic water splitting performances at elevated temperatures.

β-Lactam antibiotic detection has significant implications in food safety control, environmental monitoring and pharmacokinetics study. Here, we report the development of two BADAN-conjugated β-lactamases, E166Cb and E166Cb/N170Q, as sensitive biosensors for β-lactam antibiotic detection. These biosensors were constructed by coupling an environment-sensitive BADAN probe onto location 166 at the active site of the PenP β-lactamase E166C and E166C/N170Q mutants. They gave fluorescence turn-on signals in response to β-lactam antibiotics. Molecular dynamics simulation of E166Cb suggested that the turn-on signal might be attributed to a polarity change of the microenvironment of BADAN and the removal of the fluorescence quenching effect on BADAN exerted by a nearby Tyr-105 upon the antibiotic binding. In the detection of four β-lactams (penicillin G, penicillin V, cefotaxime and moxalactam), both E166Cb and E166Cb/N170Q delivered signal outputs in an antibiotic-concentration dependent manner with a dynamic range spanning from 10 nM to 1 μM. Compared to E166Cb, E166Cb/N170Q generally exhibited more stable signals owing to its higher deficiency in hydrolyzing the antibiotic analyte. The overall biosensor performance of E166Cb and E166Cb/N170Q was comparable to that of their respective fluorescein-modified counterparts, E166Cf and E166Cf/N170Q. But comparatively, the BADAN-conjugated enzymes showed a higher sensitivity, displayed a faster response in detecting moxalactam and a more stable fluorescence signals towards penicillin G. This study illustrates the potential of BADAN-conjugated β-lactamases as biosensing devices for β-lactam antibiotics.

Bacteria are known to be sensitive to electrical interactions with the environment. Here, the authors report on a study into how the antibacterial properties of carbon-doped titania nanotubes are affected by capacitance after charging with direct and alternating currents.

Abstract β-Lactamase inhibitory protein (BLIP), a low molecular weight protein from Streptomyces clavuligerus, has a wide range of potential applications in the fields of biotechnology and pharmaceutical industry because of its tight interaction with and potent inhibition on clinically important class A β-lactamases. To meet the demands for considerable amount of highly pure BLIP, this study aimed at developing an efficient expression system in eukaryotic Pichia pastoris (a methylotrophic yeast) for production of BLIP. With methanol induction, recombinant BLIP was overexpressed in P. pastoris X-33 and secreted into the culture medium. A high yield of ~ 300 mg/L culture secretory BLIP recovered from the culture supernatant without purification was found to be > 90% purity. The recombinant BLIP was fully active and showed an inhibition constant (K i) for TEM-1 β-lactamase (0.55 ± 0.07 nM) comparable to that of the native S. clavuligerus-expressed BLIP (0.5 nM). Yeast-produced BLIP in combination with ampicillin effectively inhibited the growth of β-lactamase-producing Gram-positive Bacillus. Our approach of expressing secretory BLIP in P. pastoris gave 71- to 1200-fold more BLIP with high purity than the other conventional methods, allowing efficient production of large amount of highly pure BLIP, which merits fundamental science studies, drug development and biotechnological applications.

The protein RpoS is responsible for mediating cell survival during the stationary phase by conferring cell resistance to various stressors and has been linked to biofilm formation. In this study, the role of the rpoS gene in Escherichia coli 0157:H7 biofilm formation and survival in water was investigated. Confocal scanning laser microscopy of biofilms established on coverslips re-vealed a nutrient-dependent role of rpoS in biofilm formation, where the biofilm biomass volume of the rpoS mutant was 2.4- to 7.5-fold the size of its rpoS+ wild-type counterpart in minimal growth medium. The enhanced biofilm formation of the rpoS mu-tant did not, however, translate to increased survival in sterile double-distilled water (ddH20), filter-sterilized lake water, or un-filtered lake water. The rpoS mutant had an overall reduction of 3.10 and 5.30 log10 in sterile ddH20 and filter-sterilized lake wa-ter, respectively, while only minor reductions of 0.53 and 0.61 log10 in viable counts were observed for the wild-type form in the two media over a 13-day period, respectively. However, the survival rates of the detached biofilm-derived rpoS+ and rpoS mutant cells were comparable. Under the competitive stress conditions of unfiltered lake water, the advantage conferred by the presence of rpoS was lost, and both the wild-type and knockout forms displayed similar declines in viable counts. These results suggest that rpoS does have an influence on both biofilm formation and survival of E. coli 0157:H7 and that the advantage conferred by rpoS is contingent on the environmental conditions. [ABSTRACT FROM AUTHOR]

The Ω-loop at the active site of β-lactamases exerts significant impact on the kinetics and substrate profile of these enzymes by forming part of the substrate binding site and posing as steric hindrance toward bulky substrates. Mutating certain residues on the Ω-loop has been a general strategy for molecular evolution of β-lactamases to expand their hydrolytic activity toward extended-spectrum antibiotics through a mechanism believed to involve enhanced structural flexibility of the Ω-loop. Yet no structural information is available that demonstrates such flexibility or its relation to substrate profile and enzyme kinetics. Here we report an engineered β-lactamase that contains an environment-sensitive fluorophore conjugated near its active site to probe the structural dynamics of the Ω-loop and to detect the binding of diverse substrates. Our results show that this engineered β-lactamase has improved binding kinetics and positive fluorescence signal toward oxyimino-cephalosporins, but shows little such effect to non-oxyimino-cephalosporins. Structural studies reveal that the Ω-loop adopts a less stabilized structure, and readily undergoes conformational change to accommodate the binding of bulky oxyimino-cephalosporins while no such change is observed for non-oxyimino-cephalosporins. Mutational studies further confirm that this substrate-induced structural change is directly responsible for the positive fluorescence signal specific to oxyimino-cephalosporins. Our data provide mechanistic evidence to support the long-standing model that the evolutionary strategy of mutating the Ω-loop leads to increased structural flexibility of this region, which in turn facilitates the binding of extended spectrum β-lactam antibiotics. The oxyimino-cephalosporin-specific fluorescence profile of our engineered β-lactamase also demonstrates the possibility of designing substrate-selective biosensing systems. [ABSTRACT FROM AUTHOR]

The fluorescein-labeled E166C mutant of the PenPC β-lactamase (E166Cf) represents a successful model in the construction of "switch-on" fluorescent biosensors from nonallosteric proteins (Chan P.-H. et al.; J. Am Chem. Soc., 2004, 126, 4074). This paper focuses on the study of the biosensing mechanism by which the E166Cf biosensor changes its fluorescence upon β-lactam binding and hydrolysis. Mass spectrometric and stopped-flow fluorescence studies of E166Cf with cefuroxime, penicillin G, and 6-aminopenicillanic acid reveal that the formation of enzyme-substrate complex enhances the fluorescence of E166Cf, and the subsequent regeneration of the free enzyme restores the weak fluorescence of E166Cf. Molecular modeling studies of E166Cf with penicillin G show that the fluorescein label is likely to share a common space with the β-lactam and thiazolidine rings of the antibiotic in the active site. This spatial clash appears to cause the fluorescein label to move from the active site to the external aqueous environment upon substrate binding and hence experience higher water exposure. Steady-state fluorescence measurements indicate that the fluorescence of E166Cf can be enhanced by 6-aminopenicillanic acid, which consists of the β-lactam and thiazolidine rings only. Thermal denaturation experiments of the wild-type enzyme, E166C, and E166Cf reveal that the E166C mutation is likely to increase the flexibility of the Ω-loop. This "modified" structural property might compensate for the possible steric effect of the fluorescein label on substrate binding. [ABSTRACT FROM AUTHOR]

The synthesis and spectroscopic properties of trans-[RuL4(C≡CAr)2] (L4 = two 1 ,2-bis- (dimethylphosphino)ethane, (dmpe)2; 1,5,9,13-tetramethyl-1 ,5,9,13-tetraazacyclohexadecane, 16-TMC; 1,- 12-dimethyl-3,4:9,10-dibenzo- 1,12-diaza-5,8-dioxacyclopentadecane, N2O2) are described. Investigations into the effects of varying the [RuL4] core, acetylide ligands, and acetylide chain length for the [-C≡C(C6H4C≡C)n-1 Ph] and [-C≡C(C6H4)n-1Ph] (n = 1-3) series upon the electronic and electrochemical characteristics of trans-[RuL4(C≡CAr)2]0/+ are presented. DFT and TD-DFT calculations have been performed on trans-[Ru(L')4(C≡CAr)2]0/+ (L' = PH3 and NH3) to examine the metal—acetylide π-interaction and the nature of the associated electronic transition(s). It was observed that (1) the relationship between the transition energy and 1/n for trans-[Ru(dmpe)2{C≡C(C6H4C≡C)n-1Ph}2] (n = 1-3) is linear, and (2) the sum of the dπRuII) → π*(C≡CAr) MLCT energy for trans-[Ru(1 6-TMC or N2O2)(C≡CAr)2] and the π- (C≡CAr) → dπRuIII) LMCT energy for trans-[Ru(16-TMC or N2O2)(C≡CAr)2]+ corresponds to the intraligand ππ* absorption energy for trans-[Ru(1 6-TMC or N2O2)(C≡CAr)2]. The crystal structure of trans-[Ru(dmpe)2- {C≡C(C6H4C≡C)2Ph}2] shows that the two edges of the molecule are separated by 41.7 Å. The electrochemical and spectroscopic properties of these complexes can be systematically tuned by modifying L4 and Ar to give E1/2 values for oxidation of trans-[RuL4(C≡CAr)2] that span over 870 mV and λmax values of trans-[RuL4(C≡CAr)2] that range from 19 230 to 31 750 cm-1. The overall experimental findings suggest that the π-back-bonding interaction in frans-[RuL4(C≡CAr)2] is weak and the [RuL4 moiety in these molecules may be considered to be playing a "dopant" role in a linear rigid π-conjugated rod. [ABSTRACT FROM AUTHOR]

The increasing emergence of new bacterial β-lactamases that can efficiently hydrolyze β-lactam antibiotics to clinically inactive carboxylic acids has created an intractable problem in the treatment of bacterial infections, and it is highly desirable to develop a useful tool that can rapidly screen bacteria for β-lactamases against a variety of antibiotic candidates in a high-throughput manner. This paper describes the use of a fluorescein-labeled β-lactamase mutant (E166Cf) as a convenient fluorescent tool to screen β-lactamases, including the Bacillus cereus β-lactamase I (PenPC), B. cereus β-lactamase II, Bacillus licheniformis PenP, Escherichia coli TEM-1, and Enterobacter cloacae P99 against various β-lactam antibiotics (penicillin G, penicillin V, ampicillin, cefuroxime, cefoxitin, moxalactam, cephaloridine), using a 96-well microplate reader. The E166Cf mutant was constructed by replacing Glu166 on the flexible Ω-loop, which is dose to the enzyme's active site, with a cysteine residue on a class A β-lactamase (B. cereus PenPC) and subsequently labeling the mutant with thiol-reactive fluorescein-5-maleimide. Such modifications significantly impaired the hydrolytic activity of the E166Cf mutant compared to that of the wild-type enzyme. The fluorescence intensity of the E166Cf mutant increases in the presence of β-lactam antibiotics. For antibiotics that are resistant to hydrolysis by the E166Cf mutant (cefuroxime, cefoxitin, moxalactam), the fluorescence signal slowly increases until it reaches a plateau. For antibiotics that can be slowly hydrolyzed by the E166Cf mutant (penicillin G, penicillin V, ampicillin), the fluorescence signal rapidly increases to the plateau and then declines after a prolonged incubation. The E166Cf mutant retains its characteristic pattern of fluorescence signals in the presence of both bacterial β-lactamases and β-lactamase-resistant antibiotics. In contrast, in the presence of both bacterial β-lactamases and β-lactamase-sensitive antibiotics, the fluorescence signals of the E166Cf mutant were decreased. The fluorescence signals from the E166Cf mutant allow an unambiguous differentiation ofβ-lactamase-resistant antibiotics from β-lactamase-sensitive ones in the screening of bacterial β-lactamases against a panel of antibiotic candidates. This simple method may provide an alternative tool in choosing potent β-lactam antibiotics for treatment of bacterial infections. [ABSTRACT FROM AUTHOR]

A luminescent molecular rectangle [Au4(μ-PAnP)2(μ-bipy)2](OTf)4 (1·(OTf)4) (PAnP = 9,10-bis-(diphenylphosphino)anthracene, bipy = 4,4'-bipyridine, X = NO3- or OTf-), synthesized from the self-assembly of the molecular "clip" Au2(μ-PAnP)(OTf)2 and bipy, shows a large rectangular cavity of 7.921 (3) × 16.76(3) Å. The electronic absorption and emission spectroscopy, and electrochemistry of the metallacyclophane, have been studied. The 14+ ions are self-assembled into 2D mosaic in the solid state via complementary edge-to-face interactions between the Ph groups. ¹H NMR titrations ratify the 1:1 complexation between 14+ and various aromatic molecules. Comparing the structures of the inclusion complexes indicates an induced-fit mechanism operating in the binding. The emission of 14+ is quenched upon the guest binding. The binding constants are determined by both ¹H NMR and fluorescence titrations. Solvophobic and ion-dipole effects are shown to be important in stabilizing the inclusion complexes. [ABSTRACT FROM AUTHOR]