We report the engineering of a new reversibly switching chromogenic protein, Dathail. Dathail was evolved from the extremely thermostable fluorescent proteins thermal green protein (TGP) and eCGP123 ...using directed evolution and ratiometric sorting. Dathail has two spectrally distinct chromogenic states with low quantum yields, corresponding to absorbance in a ground state with a maximum at 389nm, and a photo-induced metastable state with a maximum at 497nm. In contrast to all previously described photoswitchable proteins, both spectral states of Dathail are non-fluorescent. The photo-induced chromogenic state of Dathail has a lifetime of ~50min at 293K and pH7.5 as measured by UV–Vis spectrophotometry, returning to the ground state through thermal relaxation. X-ray crystallography provided structural insights supporting a change in conformation and coordination in the chromophore pocket as being responsible for Dathail's photoswitching. Neutron crystallography, carried out for the first time on a protein from the green fluorescent protein family, showed a distribution of hydrogen atoms revealing protonation of the chromophore 4-hydroxybenzyl group in the ground state. The neutron structure also supports the hypothesis that the photo-induced proton transfer from the chromophore occurs through water-mediated proton relay into the bulk solvent. Beyond its spectroscopic curiosity, Dathail has several characteristics that are improvements for applications, including low background fluorescence, large spectral separation, rapid switching time, and the ability to switch many times. Therefore, Dathail is likely to be extremely useful in the quickly developing fields of imaging and biosensors, including photochromic Förster resonance energy transfer, high-resolution microscopy, and live tracking within the cell.
Display omitted
•Dathail will be useful in imaging and biosensors, particularly pcFRET, high-resolution microscopy, and live cell tracking.•Dathail is a reversibly switching chromoprotein with two spectrally distinct chromogenic states with low quantum yields.•X-ray crystallography shows changes in conformation and coordination of the chromophore during Dathail's photoswitching.•Crystallography suggests photo-induced proton transfer from the chromophore with water-mediated proton relay to bulk solvent.•The first time that neutron crystallography has been used for a protein in the GFP family provides surprising new insights.
A method of incorporating both metal−ligand and crown ether−dialkylammonium self-assembly techniques in the efficient synthesis of metallacyclic tris2pseudorotaxanes is described. These ...multifunctional supramolecules can be prepared by the stepwise or one-pot assembly of nine separate components on account of the orthogonality of the two noncovalent assembly strategies employed. This methodology demonstrates the power of orthogonal self-assembly and may be expanded beyond the use of two recognition motifs in hopes of mimicking the complex multifunctionality of biological systems.
This is for the first time that two fluoride tolerant bacterial strains of
Bacillus spp
(KT201599 and KT201600) have been isolated from the mid-gut of third instar larvae of
Drosophila melanogaster.
...They are characterized and identified using phenotypic characters and 16S rRNA gene based molecular phylogenetic analysis. Interestingly, KT201599 and KT201600 are able to survive at 2000–2500 µg/mL sodium fluoride (NaF) concentrations. The chronic lethal concentration 50 (LC
50
) for NaF in
Drosophila
larvae is 125 µg/mL, whereas, the above-mentioned
Bacillus
strains isolated from their mid-gut are found to be more tolerant to fluoride (F) in comparison to their host. In search for biological relevance of KT201599 and KT201600, we further deciphered that these strains not only survived at high F concentrations, but also were efficient in removing the fluoride ions from the medium by 16.66 and 24.71% respectively. Thus, the study indicates that resident gut flora of
Drosophila
having the property of reducing fluoride load from the medium, might be helping the host to thrive in a fluoridated environment.
The study appraised effects of Bacillus tequilensis (KF623287), through diets or water, on the growth and health status of rohu, Labeo rohita fingerlings. A basal diet (35% protein) was fed to the ...control (without B. tequilensis supplementation) and other experimental groups in triplicate for 60 days. B. tequilensis was added either to the diets at 1 × 107 cfu/g (D1), 1 × 109 cfu/g (D2) and 1 × 1011 cfu/g (D3) or to the rearing water at 1 × 107 cfu/ml (WA1), 1 × 109 cfu/ml (WA2) and 1 × 1011 cfu/ml (WA3) every 2 days. Results indicated that group D2 had significantly (p < .05) higher weight gain (115 ± 3.25%) and improved feed utilization than other experimental groups. However, group WA3 (109 ± 3.16% weight gain) appeared as the second best after D2 group. Further, improved haematological parameters, serum lysozyme, peroxidase and superoxide anion production along with increased phagocytic and respiratory burst activities were noticed in fish that received B. tequilensis either through diets (D2) or as water additive (WA3). Increased serum IgM was noticed at day 30, while decreased thereafter. The study revealed that B. tequilensis, either through diets or as water, could improve growth, nutrient utilization and immunity in L. rohita fingerlings. However dietary administration of probiotics at a lower dose was more effective than the water additive.
•Thermodynamic performance of plain and helical microfin enhanced tubes are compared for R32 and R410A by drift flux-based entropy generation analysis.•The enhanced tube performs better below a ...critical mass flux, which increases with tube diameter and heat flux.•Critical inlet quality, below which the enhanced tube performance is superior, shifts towards lower value with an increase in mass flux.•An enhanced tube performs better above a critical operating pressure, which increases with an increase in mass flux.
In the present paper, a comparative study is carried out to assess the relative thermodynamic performance of plain and helical microfin evaporator tubes with two new emergent refrigerants, R32 and R410A. An entropy generation analysis is performed based on the drift flux model for two-phase forced convective boiling flow, and the comparison of the enhanced tube to the plain one is done quantitatively. A critical value of relevant parameters, i.e., mass flux, tube diameter, length, heat flux, inlet quality, and operating pressure, are identified, below or above which the enhanced tube is observed to be thermodynamically more favourable than the plain tube. The study reveals that by increasing the tube diameter from 8 mm to 14 mm, the critical mass flux increases from 335 kg/m2s to 495 kg/m2s for R32 and 394 kg/m2s to 565 kg/m2s for R410A, respectively, below which the enhanced tube performs better. The helical microfin tube exhibits a better performance below a critical mass flux from 412 kg/m2s to 374 kg/m2s for R32 and 492 kg/m2s to 443 kg/m2s for R410A when tube length increases from 1 m to 5 m. The critical inlet quality, below which the enhanced tube is observed to be superior, shifts to a lower value (0.29 to 0.16 for R32 and 0.44 to 0.26 for R410A), increasing the mass flux from 400 kg/m2s to 500 kg/m2s. However, the enhanced tube shows a better performance above a critical operating pressure, which increases from 11.4 bar to 16.9 bar for R32 and 9.3 bar to 13.4 bar for R410A, with increasing the mass flux from 400 kg/m2s to 500 kg/m2s. The enhanced tube performs better below a critical mass flux which increases from 336 kg/m2s to 461 kg/m2s for R32 and 399 kg/m2s to 489 kg/m2s for R410A when the heat flux is increased from 8 kW/m2 to 15 kW/m2. The overall parametric study with two emergent refrigerants enables one for effective selection of the helical microfin evaporator tube for a suitable range of relevant parameters.
The gastrointestinal (GI) microbiome in fish plays significant roles in health and disease resistance. This investigation was accomplished to enumerate, characterize and identify the potential ...probiotic bacteria from three Indian Major Carps (IMCs), viz., rohu (Labeo rohita), catla (Catla catla) and mrigal (Cirrhinus mrigala) using culture dependent methods. Altogether, 105 pathogen-inhibitory bacteria (out of 1216 isolates) were detected from three IMCs by double layer assay. 16S rRNA partial gene sequence analyses and BLAST search in the NCBI GenBank unveiled that 94.29% of the pathogen inhibitory bacteria were bacilli (99 strains) and Bacillus licheniformis by far the most common (28%). The primarily selected 27 pathogen-inhibitory strains (cumulative inhibition score ≥13) produced extracellular enzymes, while 15 of them produced all the six exo-enzymes studied (amylase, protease, lipase, cellulase, phytase and xylanase). Gut stability of the strains became apparent by their ability to grow in fish mucus and tolerance to diluted bile-juice. Finally, 14 strains were noticed as γ-hemolytic and susceptible to the commonly used antibiotics. Further, intra-peritoneal injection with γ-hemolytic strains did not induce any pathological signs or mortalities in fish, and thus were considered as safe. These 14 γ-hemolytic isolates were represented by the genus Bacillus (13) and Stenotrophomonas (1), which might form probiotic consortia for prospective use in carp culture.
The transportation system plays a crucial role in the context of socioeconomic development, whereas the highway infrastructure acts as a base for the transportation system. In recent years, a rich ...impetus has been given to the development of road infrastructure by Indian governance. There is a need to introspect how well the prevailing highway infrastructure is equipped with emergency rescue management during road accidents. Lack of ambulance service and trauma facilities along the highways results in a steady loss of lives and injuries and increases people's exposure to risks.
This study aims to determine the response time of ambulance reachability to the accident spot on Indian national highways associated with heavy commercial transportation. Also, determining the time to transport the injured to the nearest trauma facility is another factor included as an objective in this investigation.
The study adopted survey-based research, whereby the variables in the questionnaire were designed to record and assess the time for an ambulance to reach the accident spot and, from there, to transport the injured to the trauma management facility on Indian highways. Two hundred twenty-five participants who were either victims/relatives of victims or those involved in the rescue of the injured have participated in the survey. The dates of the accident events were 2017 and 2022.
The survey resulted in the identification of two categories of highway accidents. The first category of accidents happened on the highways near city limits/dense settlements, and the second category occurred on the core highways. The percentage of accidents caused on the highways either adjacent to or passing through the city limits/dense settlements was reported to be higher than the accidents on the core highways. Ninety percent of the participants reported successful contact with the ambulance call/service centre, but only ~75% success rate exists for ambulances to reach the accident scene. On the core highways, the time taken for the ambulance to arrive at the accident scene is 25-35 minutes. The results from the survey ascertained that the patients were prioritised for treatment in the nearest hospitals (irrespective of having a trauma facility) at a distance of ~12-20 km, for which the time taken is ~15-25 minutes. Importantly, from the interviews, it is understood that in many cases, these hospitals have further referred to specialty hospitals located in nearby cities or trauma centres with greater facilities. Occasions exist where the injured were taken directly to hospitals 30-40 km from the accident spot, for which the time was more than 40 minutes.
The results provide evidence that in either of the accident cases on the highways that are adjacent to/passing through the city limits or on the core highways, the total time for emergency care accessibility is nearly 60 minutes or greater; this implies that in the majority of cases, there is very meagre time left to provide emergency medical care to the needy and injured on the Indian highways to abide by the concept of golden hour. Plausible reforms backed by technology for enabling highways into 'emergency rescuable highways' are highly needed to guarantee a safer and more sustainable transportation system in India.
Transition-metal-catalyzed activation of inert C–H bonds and subsequent C–C bond formation have emerged as powerful synthetic tools for the synthesis of elaborate cyclic molecules. In this report, we ...introduce an efficient synthetic method of 3,4-unsubstituted isocoumarins adopting an electron-deficient CpERh complex as the catalyst. The use of vinylene carbonate as a vinylene transfer reagent enables the direct construction of isocoumarins from readily available benzoic acids, without any external oxidants as well as bases. The reaction mechanism is evaluated by computational analysis to find an unprecedented “rhodium shift” event within the catalytic cycle.
The formation of a desired nanostructure with concomitant patterns and functions is of utmost importance in the field of surface molecular engineering and nanotechnology. We here present a flexible ...host−guest assembly, which steers the formation of linear molecular nanostructures on surfaces by a hydrogen-bond-mediated assembly process. A linear monodendron molecular template with periodic hydrogen-bond binding sites is shown to accommodate a variety of molecules with pyridylethynyl terminals. The unit cell parameters in the transverse direction of the linear pattern can be tuned from 3.4 to 7.3 nm in response to the packing of the guest molecules with different sizes, shapes, and aggregation number. The introduction of hydrogen-bonding partners into the host template and into guest molecules is responsible for the steering of the linear pattern of guest molecules. The modular approach could greatly facilitate the ordering of guest molecules with desired functional moieties.
PDF
