The 2.1 angstrom crystal structure of copGFP, a representative member of the copepod clade within the green fluorescent protein superfamily

The green fluorescent protein (avGFP), its variants, and the closely related GFP-like proteins are characterized structurally by a cyclic tri-peptide chromophore located centrally within a conserved beta-can fold. Traditionally, these GFP family members have been isolated from the Cnidaria although recently, distantly related GFP-like proteins from the Bilateria, a sister group of the Cnidaria have been described, although no representative structure from this phylum has been reported to date. We have determined to 2.1 angstrom resolution the crystal structure of copGFP, a representative GFP-like protein from a copepod, a member of the Bilateria. The structure of copGFP revealed that, despite sharing only 19% sequence identity with GFP, the tri-peptide chromophore (Gly57-Tyr58-Gly59) of copGFP adopted a cis coplanar conformation within the conserved beta-can fold. However, the immediate environment surrounding the chromophore of copGFP was markedly atypical when compared to other members of the GFP-superfamily, with a large network of bulky residues observed to surround the chromophore. Arg87 and Glu222 (GFP numbering 96 and 222), the only two residues conserved between copGFP, GFP and GFP-like proteins are involved in autocatalytic genesis of the chromophore. Accordingly, the copGFP structure provides an alternative platform for the development of a new suite of fluorescent protein tools. Moreover, the structure suggests that the autocatalytic genesis of the chromophore is remarkably tolerant to a high degree of sequence and structural variation within the beta-can fold of the GFP superfamily. (c) 2006 Elsevier Ltd . All rights reserved.

Molecular and cellular effects of ultraviolet light-induced genotoxicity

Exposure to the solar ultraviolet spectrum that penetrates the Earth's stratosphere (UVA and UVB) causes cellular DNA damage within skin cells. This damage is elicited directly through absorption of energy (UVB), and indirectly through intermediates such as sensitizer radicals and reactive oxygen species (UVA). DNA damage is detected as strand breaks or as base lesions, the most common lesions being 8-hydroxydeoxyguanosine (8OHdG) from UVA exposure and cyclobutane pyrimidine dimers from UVB exposure. The presence of these products in the genome may cause misreading and misreplication. Cells are protected by free radical scavengers that remove potentially mutagenic radical intermediates. In addition, the glutathione-S-transferase family can catalyze the removal of epoxides and peroxides. An extensive repair capacity exists for removing (1) strand breaks, (2) small base modifications (8OHdG), and (3) bulky lesions (cyclobutane pyrimidine dimers). UV also stimulates the cell to produce early response genes that activate a cascade of signaling molecules (e.g., protein kinases) and protective enzymes (e.g., haem oxygenase). The cell cycle is restricted via p53-dependent and -independent pathways to facilitate repair processes prior to replication and division. Failure to rescue the cell from replication block will ultimately lead to cell death, and apoptosis may be induced. The implications for UV-induced genotoxicity in disease are considered.

Solubilisation of drugs within liposomal bilayers:alternatives to cholesterol as a membrane stabilising agent

Objectives The aim of this work was to investigate the effect of cholesterol on the bilayer loading of drugs and their subsequent release and to investigate fatty alcohols as an alternative bilayer stabiliser to cholesterol. Methods The loading and release rates of four low solubility drugs (diazepam, ibuprofen, midazolam and propofol) incorporated within the bilayer of multilamellar liposomes which contained a range of cholesterol (0–33 mol/mol%) or a fatty alcohol (tetradecanol, hexadecanol and octadecanol) were investigated. The molecular packing of these various systems was also investigated in Langmuir monolayer studies. Key findings Loading and release of drugs within the liposome bilayer was shown to be influenced by their cholesterol content: increasing cholesterol content was shown to reduce drug incorporation and inclusion of cholesterol in the bilayer changed the release profile of propofol from zero-order, for phosphatidyl choline only liposomes, to a first-order model when 11 to 33 total molar % of cholesterol was present in the formulation. At higher bilayer concentrations substitution of cholesterol with tetradecanol was shown to have less of a detrimental impact on bilayer drug loading. However, the presence of cholesterol within the liposome bilayer was shown to reduce drug release compared with fatty alcohols. Monolayer studies undertaken showed that effective mean area per molecule for a 1,2-distearoyl-sn-glycero-3-phosphocholine (DSPC) : cholesterol mixture deviated by 9% from the predicted area compared with 5% with a similar DSPC : tetradecanol mixture. This evidence, combined with cholesterol being a much more bulky structure, indicated that the condensing influence of tetradecanol was less compared with cholesterol, thus supporting the reduced impact of tetradecanol on drug loading and drug retention. Conclusions Liposomes can be effectively formulated using fatty alcohols as an alternative bilayer stabiliser to cholesterol. The general similarities in the characteristics of liposomes containing fatty alcohols or cholesterol suggest a common behavioural influence for both compounds within the bilayer.

Premium e-grocery:exploring value in logistics integrated service solutions

E-grocery is gradually becoming viable or a necessity for many families. Yet, most e-supermarkets are seen as providers of low value "staple" and bulky goods mainly. While each store has a large number of SKU available, these products are mainly necessity goods with low marginal value for hedonistic consumption. A need to acquire diverse products (e.g., organic), premium priced products (e.g., wine) for special occasions (e.g., anniversary, birthday), or products just for health related reasons (e.g., allergies, diabetes) are yet to be served via one-stop e-tailers. In this paper, we design a mathematical model that takes into account consumers' geo-demographics and multi-product sourcing capacity for creating critical mass and profit. Our mathematical model is a variant of Capacitated Vehicle Routing Problem with Time Windows (CVRPTW), which we extend by adding intermediate locations for trucks to meet and exchange goods. We illustrate our model for the city of Istanbul using GIS maps, and discuss its various extensions as well as managerial implications.

The automation of resonant thermometer probe measurements

An ultrasonic thermometer has been developed for high temperature measurement over a wide temperature range. It is particularly suitable for use in measuring nuclear fuel rod centerline temperatures in advanced liquid metal and high flux nuclear reactors. The thermometer which was designed to determine fuel temperature up to the fuel melting point, utilizes the temperature dependence of the ultrasonic propagation velocity (related to the elastic modulus} in a thin rod sensor as the temperature transducing mechanism. A pulse excitation technique has been used, where the mechanical resonator at the remote end of the acoustic·line is madto vibrate. Its natural frequency is proportional to the ultrasonic velocity in the material. This is measured by the electronic instrumentation and enables a frequency­ temperature or period-temperature calibration to be obtained. A completely digital automatic instrument has been designed, constructed and tested to track the resonance frequency of the temperature sensors. It operates smoothly over a frequency range of about 30%, more than the maximum working range of most probe materials. The control uses the basic property of a resonator that the stored energy decays exponentially at the natural frequency of the resonator.The operation of the electronic system is based on a digital multichannel transmitter that is capable of operating with a predefined number of cycles in the burst. this overcomes a basic defect in the previous deslgn where the analogue time-delayed circuits failed to hold synchronization and hence automatic control could be lost. Development of a particular type of temperature probe, that is small enough to fit into a standard 2 mm reactor tube has made the ultrasonic thermometer a practicable device for measuring fuel temperature. The bulkiness of previous probes has been overcome, the new design consists of a tuning fork, integral with a 1mm line, while maintaining a frequency of no more than 100 kHz. A magnetostrictive rod, acoustically matched to the probe is used to launch and receive the acoustic oscillations. This requires a magnetic bias and the previously used bulky magnets have been replaced by a direct current coil. The probe is supported by terminating the launcher with a short heavy isolating rod which can be secured to the reactor structure. This support, the bias and launching coil and the launcher are made up into a single compact unit. On the material side an extensive study of a wide range of refractory materials identified molybdenum, iridium, rhenium and tungsten as satisfactory for a number of applications but mostly exhibiting to some degree a calibration drift with thermal cycling. When attention was directed to ceramic materials, Sapphire (single crystal alumina) was found to have numerous advantages, particularly in respect of stability of calibration which remained with ±2°C after many cycles to 1800oC. Tungsten and thoriated tungsten (W - 2% Tho2) were also found to be quite satisfactory to 1600oC, the specification for a Euratom application.

The synthesis of novel biodegradable polyesters and copolyesters from anhydrosulfites

This thesis was concerned primarily with the synthesis and the ring-opening polymerisation of anhydrosulfites (1,3,2-dioxa-thiolan-4-one-2-oxides), and secondly with the copolymerisation of anhydrosulfites with -caprolactone. The polyesters and copolyesters synthesised are of considerable interest in medical applications and also for use as environmental friendly packaging. A range of anhydrosulfites were prepared according to an established method. Aliphatic anhydrosulfites were obtained with a level of purity satisfactory for polymerisation whereas aromatic anhydrosulfites decomposed during distillation and purification by chromatographic techniques. Aliphatic anhydrosulfites with a substituent, such as methyl, isopropyl, n-butyl and isobutyl were studied by NMR spectroscopy. Analysis of these spectra revealed that the five-membered anhydrosulfite ring was puckered and that when the substituent was bulky, rotations about the alkyl chains were restricted. A wide range of anionic initiators may be used to initiate anhydrosulfites. Lithium alkyls turned out to be more successful than alkali metal alkoxides and amides. The molecular weights were found to depend on the basicity of the initiator, the monomer-to-initiator ratio, the nature of the solvent and the polymerisation temperature. The molecular weight M0 of poly(L-lactic acid) ranged from (0.5 to 6)x104. Highly crystalline and purely isotactic poly(lactic acid) was synthesised from L-lactic acid anhydrosulfite (L-LAAS) whereas DL-LAAS led to an amorphous polymer with randomly distributed D-and L-lactic units. This indicated that this polymerisation was not stereoselective. However, the bulkiness of the substituent in the anhydrosulfites molecule was found to influence the stereoselectivity of the polymerisation, thus polyesters with isobutyl or n-butyl pendant group were preferentially isotactic. Block-copolymers of ε-caprolactone and several anhydrosulfites were successfully produced. Block-copolymers of LAAS with ε-caprolactone were also synthesised, but the incorporation of caprolactone units was rather small. In contrast, random copolymerisation of LAAS and ε-caprolactone led to polymers with blocky structures similar to those obtained in the block-copolymerisation of LAAS with  ε-caprolactone.

Dipole moments and Kerr effect of poly(n-vinylcarbazole) and its complexes with trinitrofluorenone

N-vinylcarbazole was polymerised using the free radical catalyst (azo-bisisobutyronitrile) and cationic catalysts (boron-trifluoride etherate and aluminium chloride). The polymers produced were characterised by molecular weight measurements and powder x-ray diffraction. The tacticity of the polymer samples was determined using proton and carbon-13 nuclear magnetic resonance spectroscopy. Measurements of their static dielectric permittivity and electro-optical birefringence (Kerr effect) in solution in 1,4-dioxane were carried out over a range of temperatures. The magnitudes of the dipole moments and Kerr constants were found to vary with changes in the tacticity of poly(N-vinylcarbazole). The results of these measurements support the view that the stereostructure of poly(N-vinylcarbazole) is sensitive to the mechanism of polymerisation. These results, together with proton and carbon-13 N.M.R. data, are discussed in terms of the possible conformations of the polymer chains and the relative orientation of the bulky carbazole side groups. The dielectric and molecular Kerr effect studies have also been carried out on complexes formed between 2,4,7-trinitro-9-fluorenone (TNF) and different stereoregular forms of poly(N-vinylcarbazole) in solution in 1,4-dioxane. The differences in the molar Kerr constants between pure (uncomplexed) and complexed poly(N-vinylcarbazole) samples were attributed to changes in optical anisotropy and dipole moments. A molecular modelling computer program Desktop Molecular Modeller was used to examine the 3/1 helical isotactic and 2/1 helical syndiotactic forms of poly(N-vinylcarbazole). These models were used to calculate the pitch distances of helices and the results were interpreted in terms of van der Waal's radii on TNF. This study indicated that the pitch distance in 3/1 isotactic helices was large enough to accommodate the bulky TNF molecules to form sandwich type charge transfer complexes whereas the pitch distance in syndiotactic poly(N-vinylcarbazole) was smaller and would not allow a similar type of complex formation.

Investigation of some critical aspects of on-line surface measurement by a wavelength-division-multiplexing technique

Some critical aspects of a new kind of on-line measurement technique for micro and nanoscale surface measurements are described. This attempts to use spatial light-wave scanning to replace mechanical stylus scanning, and an optical fibre interferometer to replace optically bulky interferometers for measuring the surfaces. The basic principle is based on measuring the phase shift of a reflected optical signal. Wavelength-division-multiplexing and fibre Bragg grating techniques are used to carry out wavelength-to-field transformation and phase-to-depth detection, allowing a large dynamic measurement ratio (range/resolution) and high signal-to-noise ratio with remote access. In effect the paper consists of two parts: multiplexed fibre interferometry and remote on-machine surface detection sensor (an optical dispersive probe). This paper aims to investigate the metrology properties of a multiplexed fibre interferometer and to verify its feasibility by both theoretical and experimental studies. Two types of optical probes, using a dispersive prism and a blazed grating, respectively, are introduced to realize wavelength-to-spatial scanning.

Antitumour imidazotetrazines: probes for the major groove of DNA

The antitumour imidazotetrazinones are believed to act as prodrugs for the triazene series of alkylating agents, showing a marked pteference for the alkylation of the middle guanine residue in a run of three or more contiguous guanines. However, the. exact nature of the interactions of imidazotetrazinones within the micro~environment of DNA are; as yet unknown. In order to examine such interactions a three pronged approach involving molecular modelling, synthetic chemistry and biological analysis has been undertaken during the course of this project. . Molecular modelling studies have shown that for the 8-carboxamido substituted imidazotetrazinones antitumour activity is dependent upon the. presence of a free NH group which can be involved in the formation of both intramolecular and intermolecular hydrogen bonds, and the presence of a non-bulky substituent with a small negative potential . volume. Modelling studies involving the docking of .mitozolomide into the major groove of DNA in the region of a triguanine sequence has shown that a number of hydrogen bonding interactions are feasible. A series of 8-substituted carboxamide derivatives of mitozolomide have been synthesised via the 8-acid chloride and 8-carboxylic acid derivatives including a number of peptide analogues. The peptide derivatives were based upon the key structural features of the helix-turn-helix motif of DNA-binding proteins with a view to developing agents that are capable of binding to DNA with greater selectivity. An examination of the importance of intramolecular hydrogen bonding in influencing the antitumour activity:of :the imidazotetrazinones has led to the synthesis of the novel pyrimido[4',5' :4,3]pyrazolo[5,1-d]-1,2,3,5-tetrazine ring system. In general, in vitro cytotoxicity assays showed that the new derivatives were less active against the TLX5 lymphoma cell line. than the parent compound mitozolomide despite an increased potential for hydrogen bonding interactions. Due to the high reactivity of the: tetrazinone ring system it is difficult to study the interactions between the imidazotetrazinones and DNA. Consequently a number of structural analogues that are stable under physiological conditions have been. prepared based upon the 1,2,3 triazin-4(3H)-one ring system fused with both benzene and pyrazole rings. Although the 3-methylbenzotriazinones failed to antagonise the cytotoxic activity of temozolomide encouraging results with a 3-methylpyrazolotriazinone may suggest the existence of an imidazotetrazinone receptor site within DNA. The potential of guanine rich sequences to promote the alkylating selectivity of imidazotetrazinones by acting as a catalyst for ring cleavage and thereby generation of the alkylating agent was examined. Experiments involving the monitoring: of the rate of breakdown of mitozolomide incubated in the presence of synthetic oIigonucleotides did not reveal any catalytic effect resulting from the DNA. However, it was noted that the breakdown of mitozolomide was dependent upon the type of buffer used in the incubations and this may indeed mask any catalysis by the oligonucleotides.

Ultra small integrated optical fiber sensing system

This paper introduces a revolutionary way to interrogate optical fiber sensors based on fiber Bragg gratings (FBGs) and to integrate the necessary driving optoelectronic components with the sensor elements. Low-cost optoelectronic chips are used to interrogate the optical fibers, creating a portable dynamic sensing system as an alternative for the traditionally bulky and expensive fiber sensor interrogation units. The possibility to embed these laser and detector chips is demonstrated resulting in an ultra thin flexible optoelectronic package of only 40 µm, provided with an integrated planar fiber pigtail. The result is a fully embedded flexible sensing system with a thickness of only 1 mm, based on a single Vertical-Cavity Surface-Emitting Laser (VCSEL), fiber sensor and photodetector chip. Temperature, strain and electrodynamic shaking tests have been performed on our system, not limited to static read-out measurements but dynamically reconstructing full spectral information datasets.

Investigation of some critical aspects of on-line surface measurement by a wavelength-division-multiplexing technique

Some critical aspects of a new kind of on-line measurement technique for micro and nanoscale surface measurements are described. This attempts to use spatial light-wave scanning to replace mechanical stylus scanning, and an optical fibre interferometer to replace optically bulky interferometers for measuring the surfaces. The basic principle is based on measuring the phase shift of a reflected optical signal. Wavelength-division-multiplexing and fibre Bragg grating techniques are used to carry out wavelength-to-field transformation and phase-to-depth detection, allowing a large dynamic measurement ratio (range/resolution) and high signal-to-noise ratio with remote access. In effect the paper consists of two parts: multiplexed fibre interferometry and remote on-machine surface detection sensor (an optical dispersive probe). This paper aims to investigate the metrology properties of a multiplexed fibre interferometer and to verify its feasibility by both theoretical and experimental studies. Two types of optical probes, using a dispersive prism and a blazed grating, respectively, are introduced to realize wavelength-to-spatial scanning.

Ultra small integrated optical fiber sensing system

This paper introduces a revolutionary way to interrogate optical fiber sensors based on fiber Bragg gratings (FBGs) and to integrate the necessary driving optoelectronic components with the sensor elements. Low-cost optoelectronic chips are used to interrogate the optical fibers, creating a portable dynamic sensing system as an alternative for the traditionally bulky and expensive fiber sensor interrogation units. The possibility to embed these laser and detector chips is demonstrated resulting in an ultra thin flexible optoelectronic package of only 40 µm, provided with an integrated planar fiber pigtail. The result is a fully embedded flexible sensing system with a thickness of only 1 mm, based on a single Vertical-Cavity Surface-Emitting Laser (VCSEL), fiber sensor and photodetector chip. Temperature, strain and electrodynamic shaking tests have been performed on our system, not limited to static read-out measurements but dynamically reconstructing full spectral information datasets.

Blue-to-red tunable SHG from a diode-pumped PPKTP waveguide

Summary form only given. Broadly tunable compact visible laser sources in the spectral region of 500-650 nm are valuable in biophotonics, photomedicine and for many applications including spectroscopy, laser projection and confocal microscopy. Unfortunately, commercially available lasers of this spectral range are in practice bulky and inconvenient in use. An attractive method for the realization of portable visible laser sources is the frequency-doubling of the infrared laser diodes in a nonlinear crystal containing a waveguide [1]. Nonlinear crystal waveguides that offer an order-of-magnitude increase in the IR-to-visible conversion efficiency also enable a very different approach to second-harmonic generation (SHG) tunability in periodically-poled crystals, promising order-of-magnitude increase of wavelength range for SHG conversion. This is possible by utilization of a significant difference in the effective refractive indices of the high-order and low-order modes in multimode waveguides [2]. The recent availability of low-cost, good quality semiconductor diode lasers, offering the coverage of a broad spectral range between 1 µ?? and 1.3 µp? [3,4], in combination with well-established techniques to fabricate good quality waveguides in nonlinear crystals, allows compact tunable CW laser sources in the visible spectral region to be realized [2].

Highly efficient SHG at 561 nm using a QD laser and a PPLN waveguide

Compact CW lasers in the visible spectral region are of great importance for vast number of applications including biophotonics, photomedicine, spectroscopy and confocal microscopy. Currently, commercially available lasers of this spectral region are bulky, expensive and inconvenient in use. Also, there is a lack of diode lasers emitting in the visible spectral range, particularly in the yellow region, where a range of important fluorescent probes are optimally excited. An attractive way to realize a compact yellow laser source is second harmonic generation (SHG) in a periodically poled nonlinear crystal containing a waveguide which allows high-efficient frequency conversion even at moderate power level. In this respect, periodically poled lithium niobate (PPLN) waveguided crystal is one of the best candidates for efficient SHG. In recent years, the progress made with the fabrication of good quality waveguides in PPLN crystals in combination with availability of low-cost, good quality semiconductor diode lasers, offering the coverage of a broad spectral range between 1 µm and 1.3 µm, allows compact CW laser sources in the visible spectral region to be realized.

Recent developments in heterogeneous catalysis for the sustainable production of biodiesel

The quest for energy security and widespread acceptance of the anthropogenic origin of rising CO2 emissions and associated climate change from combusting fossil derived carbon sources, is driving academic and commercial research into new routes to sustainable fuels to meet the demands of a rapidly rising global population. Biodiesel is one of the most readily implemented and low cost, alternative source of transportation fuels to meet future societal demands. However, current practises to produce biodiesel via transesterification employing homogeneous acids and bases result in costly fuel purification processes and undesired pollution. Life-cycle calculations on biodiesel synthesis from soybean feedstock show that the single most energy intensive step is the catalytic conversion of TAGs into biodiesel, accounting for 87% of the total primary energy input, which largely arises from the quench and separation steps. The development of solid acid and base catalysts that respectively remove undesired free fatty acid (FFA) impurities, and transform naturally occurring triglycerides found within plant oils into clean biodiesel would be desirable to improve process efficiency. However, the microporous nature of many conventional catalysts limits their ability to convert bulky and viscous feeds typical of plant or algal oils. Here we describe how improved catalyst performance, and overall process efficiency can result from a combination of new synthetic materials based upon templated solid acids and bases with hierarchical structures, tailored surface properties and use of intensified process allowing continuous operation.