Over last decades, several studies have been focused on short-term high light stress in lichens under laboratory conditions. Such studies reported a strong photoinhibition of photosynthesis accompanied by a partial photodestruction of PSII, involvement of photoprotective mechanisms, and resynthetic processes into gradual recovery. In our paper, we applied medium [800 µmol(photon) m -2 s -1 ] light stress to induce negative changes in PSII funcioning as well as pigment and glutathione (GSH) content in two Antarctic fruticose lichen species. Chlorophyll (Chl) fluorescence parameters, such as potential and effective quantum yield of photosynthetic processes and fast transients (OJIP) recorded during high light exposition and recovery, revealed that Usnea antarctica was less susceptible to photoinhibition than U. aurantiaco-atra. This might be supported by a more pronounced high light-induced reduction in Chl a and b contents in U. aurantiaco-atra compared with U. antarctica. In both experimental species, total GSH showed an initial increase during the first 30-40 min of high light treatment followed by a decrease (60 min) and an increase during dark recovery. Full GSH recovery, however, was not finished in U. aurantiaco-atra even after 5 h indicating lower capacity of photoprotective mechanisms in the species. OJIP curves showed high light-induced decrease in both species, however, the recovery of the OJIPs shape to pre-photoinhibitory values was faster and more apparent in U. antarctica than in U. aurantiaco-atra. The results are discussed in terms of sensitivity of the two species to photoinhibition and their photosynthetic performance in natural environment.
The human antimicrobial peptide LL-37 is a cationic peptide with antimicrobial activity against both Gram-positive and Gram-negative microorganisms. This work describes the development of an expression system based on Escherichia coli capable of high production of the recombinant LL-37. The fusion protein Trx-LL-37 was expressed under control of T7 promoter. The expression of T7 polymerase in the E. coli strain constructed in this work was controlled by regulation mechanisms of the arabinose promoter. The expression plasmid was stabilized by the presence of parB locus which ensured higher homology of the culture during cultivation without antibiotic selection pressure. This system was capable of producing up to 1 g of fusion protein per 1 l of culture. The subsequent semipreparative HPLC allowed us to isolate 40 mg of pure LL-37. LL-37 showed high antimicrobial activity against both Gram-negative and Gram-positive microorganisms. Its activity against Candida albicans was practically nonexistent. Minimal Inhibition Concentration (MIC) determined for E. coli was 1.65 microM; for Staphylococcus aureus 2.31 microM, and for Enterococcus faecalis 5.54 microM. The effects of cathelicidin on E. coli included the ability to permeabilize both cell membranes, as could be observed by the increase of beta-galactosidase activity in extracellular space in time. Physiological changes were studied by scanning electron microscopy; Gram-positive microorganisms did not show any visible changes in cell shapes while the changes observed on E. coli cells were evident. The results of this work show that the herein designed expression system is capable of producing adequate quantities of active human antimicrobial peptide LL-37.
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