Metabolic engineering of ketocarotenoids biosynthetic pathway in Chlamydomonas reinhardtii strain CC-4102 nam trung tran * & Ralf Kaldenhoff in Chlamydomonas reinhardtii, ketocarotenoid biosynthesis is limited to the diploid zygospore stage. in this study, we attempted to engineer the ketocarotenoid pathway into Chlamydomonas haploid vegetative green cells by overexpressing the key enzyme ß-carotene ketolase (crBKt). We chose strain CC-4102 for the approach; competitive pathways, α-carotene biosynthesis and xanthophyll cycle are silenced in this strain. Driven by the strong constitutive HSP70/RBCS2 promoter CrBKT overexpression resulted in the production of canthaxanthin, the ketolation product from ß-carotene as well as a drastic reduction in the chlorophyll concentration. intriguingly, these phenotypes could only be detected from lines transformed and grown heterotrophically in the dark. once exposed to light, these transformants lost the aforementioned phenotypes as well as their antibiotic resistance. this phenomenon is in agreement with the fact that we were unable to recover any canthaxanthinproducing line among light-selected transformants. Ketocarotenoids are a special group of carotenoids characterized by the presence of one or several carbonyl groups in their ß-ionone rings. The electron-withdrawing effect of keto moieties on the carotenoid's conjugated π-system results in a slight shift of absorption maximum from 450 nm (yellow) to 470 nm (red). Thus, ketocarotenoids are easily recognizable by their red hue. Due to their excellent anti-oxidative characteristics, ketocarotenoids such as astaxanthin and canthaxanthin rank among the highest-value products on the carotenoid pigment market 1. While market demands are still being met mainly by the chemical industry, the interests in ketocarotenoids produced in biological systems has witnessed skyrocketing growth in the recent years 2. In contrast to non-ketolated carotenoids such as ß-carotene, lutein, zeaxanthin, violaxanthin etc., all of which are ubiquitous among photosynthetic organisms, the biosynthesis of ketocarotenoids is limited to a small groups of organisms including several marine bacteria 3 , the fungus Xanthophyllomyces dendrorhous 4 , microalgae 5 and flowers of Adonis aestivalis 6. Most prominent among these species is the unicellular green alga Haematococcus pluvialis, which can accumulate astaxanthin at very high levels up to 7% of its dried weight 7. Huge algal farms growing Haematococcus for astaxanthin extraction have been established in several countries including the USA, Israel and China 8. Industrial production of astaxanthin in Haematococcus, however is hampered by several limiting factors such as the alga's slow growth rate and low cell density, high risk of contamination 9 , parasitic disease 10 , high energy cost for induction of carotenogenesis (requiring high light intensity) and difficulty breaking the thick cell wall of haematocysts 11. Thus, there is a great and obvious need to develope new, alternative biological platforms ...
Aquaporins are important and well-studied water channel membrane proteins. However, being membrane proteins, sample preparation for functional analysis is tedious and time-consuming. In this paper, we report a new approach for the co-translational insertion of two aquaporins from Escherichia coli and Nicotiana tabacum using the CFPS system. This was done in the presence of liposomes with a modified procedure to form homogenous proteo-liposomes suitable for functional analysis of water permeability using stopped-flow spectrophotometry. Two model aquaporins, AqpZ and NtPIP2;1, were successfully incorporated into the liposome in their active forms. Shifted green fluorescent protein was fused to the C-terminal part of AqpZ to monitor its insertion and status in the lipid environment. This new fast approach offers a fast and straightforward method for the functional analysis of aquaporins in both prokaryotic and eukaryotic organisms.
In this work, the adsorptive removal of methylene blue and methyl orange by UiO-66-NO2 nanoparticles was studied. The influence of pH on the adsorption capacity was assessed. The kinetics of the adsorption process were investigated and compared with pseudo-first-order, pseudo-second-order, Elovich, and intraparticle models. The kinetics of the adsorption fits moderately with the pseudo-first-order, but perfectly fits with pseudo-second-order models, and has a very good fit with the Elovich and intraparticle models. The adsorption isotherms were measured and compared with the Langmuir and Freundlich models. The adsorption capacity of methyl orange (MO) on UiO-66-NO2 nanoparticles (142.9 mg/g) was over three times higher than that of methylene blue (MB) on the nanoparticles (41.7 mg/g). The discrepancy between these capacities was attributed to the presence of the -NO2 functional group, which caused a strong negative mesomeric effect in the metal-organic framework structure.
The increased use of wireless technology causes a significant exposure increase for all living organisms to radio frequency electromagnetic fields (RF-EMF). This comprises bacteria, animals, and also plants. Unfortunately, our understanding of how RF-EMF influences plants and plant physiology remains inadequate. In this study, we examined the effects of RF-EMF radiation on lettuce plants (Lactuca sativa) in both indoor and outdoor environments using the frequency ranges of 1890–1900 MHz (DECT) at 2.4 GHz and 5 GHz (Wi-Fi). Under greenhouse conditions, RF-EMF exposure had only a minor impact on fast chlorophyll fluorescence kinetics and no effect on plant flowering time. In contrast, lettuce plants exposed to RF-EMF in the field showed a significant and systemic decrease in photosynthetic efficiency and accelerated flowering time compared to the control groups. Gene expression analysis revealed significant down-regulation of two stress-related genes in RF-EMF-exposed plants: violaxanthin de-epoxidase (VDE) and zeaxanthin epoxidase (ZEP). RF-EMF-exposed plants had lower Photosystem II’s maximal photochemical quantum yield (FV/FM) and non-photochemical quenching (NPQ) than control plants under light stress conditions. In summary, our results imply that RF-EMF might interfere with plant stress responses and reduced plant stress tolerance.
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