• Journal of Microbiology and Biotechnology
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• v.32 no.8
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• pp.989-1002
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• 2022

Cephalopods, in particular octopus (Octopus vulgaris), have the ability to alter their appearance or body pattern by showing a wide range of camouflage by virtue of their chromatophores, which contain nanostructured granules of ommochrome pigments. Recently, the antioxidant and antimicrobial activities of ommochromes have become of great interest; therefore, in this study, the pH-dependent redox effect of the extraction solvent on the antioxidant potential and the structural characterization of the pigments were evaluated. Cell viability was determined by the microdilution method in broth by turbidity, MTT, resazurin, as well as fluorescence microscopy kit assays. A Live/Dead Double Staining Kit and an ROS Kit were used to elucidate the possible inhibitory mechanisms of ommochromes against bacterial and fungal strains. The results obtained revealed that the redox state alters the color changes of the ommochromes and is dependent on the pH in the extraction solvent. Natural phenoxazinone (ommochromes) is moderately toxic to the pathogens Staphylococcus aureus, Bacillus subtilis, Salmonella Typhimurium and Candida albicans, while the species Pseudomonas aeruginosa and Pseudomonas fluorescens, and the filamentous fungi Aspergillus parasiticus, Alternaria spp. and Fusarium verticillioides, were tolerant to these pigments. UV/visible spectral scanning and Fourier- transform infrared spectroscopy (FTIR) suggest the presence of reduced ommatin in methanol/ HCl extract with high intrinsic fluorescence.

• Proceedings of the Korean Biophysical Society Conference
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• 1996.07a
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• pp.13-13
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• 1996

Aggregation of human $\alpha$$_1$-antitrypsin ($\alpha$$_1$-AT) during folding occurs both in vitro and in vivo. In vivo aggregates of mutant $\alpha$$_1$-AT such as $M_{malton}$ (Phe52 deleted) and Z (Glu342 longrightarrowLys) variants have pathological consequences. In order to analyze the process of $\alpha$$_1$-AT aggregation in detail, the folding-unfolding kinetics of $\alpha$$_1$-AT was examined by monitoring intrinsic Trp fluorescence and ANS binding. (omitted)

• Applied Chemistry for Engineering
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• v.31 no.2
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• pp.220-225
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• 2020

Hair is made of proteins containing various amino acids. Ultraviolet (UV) radiation is believed to be responsible for the most damaging effects of sunlight, and also plays an important role in hair aging. The purpose of this study was to investigate the changes in morphological and chemical structures after ultraviolet B (UVB) irradiation of human hair. The UVB-irradiated hair showed characteristic morphological and structural changes, compared to those of the normal hair. The result from a scanning electron microscope (SEM) equipped with an energy dispersive X-ray diffractometer (EDX) showed that the scale of UV-irradiated hair appeared to be rough and the amount of oxygen element was higher than that of the normal hair. Fluorescence and three dimensional (3D) topographical images were obtained by a confocal laser scanning microscope (CLSM). In 3D images, the green emission intensity of normal hair was much higher than that of fluorescing UVB-irradiated hair. The intensity of green emission reflects the intrinsic fluorescence of hair protein. Also, a fluorescent imaging method using fluorescamine reagent was used to identify the free amino groups resulting from a peptide bond breakage in UVB-irradiated hair. Strong blue fluorescence of UVB-irradiated hair, which indicates a very high level of amino groups, was observed by CLSM. Therefore, the fluorescamine as an extrinsic fluorescence could provide a useful tool to identify the peptide bond breakage in UVB-irradiated hair. Infrared image mapping was also employed to assess the cross-sections of normal and UVB-irradiated specimens to examine the oxidation of disulfide bonds. The degree of peak areas with strong absorbance for the disulfide mono-oxide was spread from the outside to the inside of hair. The spectroscopic techniques used alone, or in combination, launch new possibilities in the field of hair cosmetics.

• Archives of Pharmacal Research
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• v.21 no.6
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• pp.677-682
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• 1998

The present study was designed to examine the metabolism of 1-anilino-8-naphthalene sulfonate (ANS), an anionic compound which is transported into liver via "multispecific organ ic anion transporter", with rat hepatic microsomes. TLC analysis indicated that the fluorescent metabolites were not produced to a measurable extent, which made it possible to assess the ANS metabolism by measuring the fluorescence disappearance. The metabolism of ANS was remarkably inhibited by the presence of SKF-525A as well as by the substitution of 02 by CO gas. ANS metabolism by microsomes also required NADPH as a cofactor. These results indicated that the microsomal monooxygenase system might be mainly responsible for the ANS metabolism. The maximum velocity ($V_{max}$) and Michaelis constant ($K_m$) were calculated to be $4.3{\pm}0.2$ nmol/min/mg protein and $42.1{\pm}2.0\;{\mu}M$, respectively. Assuming that 1g of liver contains 32mg of microsomal protein, the $V_{max}$ value was extrapolated to that per g of liver ($V_{max}^I$). The intrinsic metabolic clearance ($CL_{int}$) under linear conditions calculated from this in vitro metabolic study was 3.3ml/min/g liver, being comparable with that (3.0ml/min/g liver) calculated by analyzing the in vivo plasma disappearance curve in a previous study. Furthermore, the effects of other organic anions on the metabolism of ANS were examined. Bromophenolblue (BPB) and rose bengal (RB) competitively inhibited the metabolism of ANS, while BSP inhibited it only slightly. The inhibition constant ($K_i$) of BPB ($6\;{\mu}M$) was much smaller than that of RB ($200\;{\mu}M$). In conclusion, the microsomal monooxygenase system plays a major role in the metabolism of ANS, and other unmetabolizable organic anions (BPB and RB) compete for this metabolism.

• Asian Pacific Journal of Cancer Prevention
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• v.14 no.4
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• pp.2637-2641
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• 2013

6,8-Dihydroxy-7-methoxy-1-methyl-azafluorenone (DMMA), a purified compound from Polyalthia cerasoides roots, is cytotoxic to various cancer cell lines. The aims of this study were to demonstrate the type of cancer cell death and the mechanism(s) involved. DMMA inhibited cell growth and induced apoptotic death in human leukemic cells (HL-60, U937, MOLT-4), human breast cancer MDA-MB231 cells and human hepatocellular carcinoma HepG2 cells in a dose dependent manner, with $IC_{50}$ values ranging between 20-55 ${\mu}M$. DMMA also decreased cell viability of human peripheral blood mononuclear cells. The morphology of cancer cells induced by the compound after staining with propidium iodide and examined under a fluorescence microscope was condensed nuclei and apoptotic bodies. Mitochondrial transmembrane potential (MTP) was decreased after 24h exposure in all five types of cancer cells. DMMA-induced caspase-3, -8, and -9 activity was strongly induced in human leukemic HL-60 and MOLT-4 cells, while in U937-, MDA-MB231- and HepG2-treated cells there was partial induction of caspase. In conclusion, DMMA-induced activation of caspase-8 and -9 resulted in execution of apoptotic cell death in human leukemic HL-60 and MOLT-4 cell lines via extrinsic and intrinsic pathways.

• The Korean Journal of Physiology and Pharmacology
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• v.4 no.1
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• pp.63-72
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• 2000

Chronic exposure to cadmium (Cd) results in an inhibition of protein endocytosis in the renal proximal tubule, leading to proteinuria. In order to gain insight into the mechanism by which Cd impairs the protein endocytosis, we investigated the effect of Cd on the acidification of renal cortical endocytotic vesicles (endosomes). The endosomal acidification was assessed by measuring the pH gradient-dependent fluorescence change, using acridine orange or FITC-dextran as a probe. In renal endosomes isolated from Cd-intoxicated rats, the $V_{max}$ of ATP-driven fluorescence quenching ($H^+-ATPase$ dependent intravesicular acidification) was significantly attenuated with no substantial changes in the apparent $K_m,$ indicating that the capacity of acidification was reduced. When endosomes from normal animals were directly exposed to free Cd in vitro, the $V_{max}$ was slightly reduced, whereas the $K_m$ was markedly increased, implying that the biochemical property of the $H^+-ATPase$ was altered by Cd. In endosomes exposed to free Cd in vitro, the rate of dissipation of the transmembrane pH gradient after $H^+-ATPase$ inhibition appeared to be significantly faster compared to that in normal endosomes, indicating that the $H^+-conductance$ of the membrane was increased by Cd. These results suggest that in long-term Cd-exposed animals, free Cd ions liberated in the proximal tubular cytoplasm by lysosomal degradation of cadmium-metallothionein complex (CdMT) may impair endosomal acidification 1) by reducing the $H^+-ATPase$ density in the endosomal membrane, 2) by suppressing the intrinsic $H^+-ATPase$ activity, and 3) possibly by increasing the membrane conductance to $H^+$ ion. Such effects of Cd could be responsible for the alterations of proximal tubular endocytotic activities, protein reabsorption and various transporter distributions observed in Cd-exposed cells and animals.

• Molecules and Cells
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• v.27 no.6
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• pp.681-687
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• 2009

We examined the effects of synthetic signal peptides, wild-type (WT) and export-defective mutant (MT) of ribose-binding protein, on the conformational changes of signal recognition particle 54 homologue (Ffh) in Escherichia coli. Upon interaction of Ffh with WT peptide, the intrinsic Tyr fluorescence, the transition temperature of thermal unfolding, and the GTPase activity of Ffh decreased in a peptide concentration-dependent manner, while the emission intensity of 8-anilinonaphthalene-1-sulfonic acid increased. In contrast, the secondary structure of the protein was not affected. Additionally, polarization of fluorescein-labeled WT increased upon association with Ffh. These results suggest that WT peptide induces the unfolded states of Ffh. The WT-mediated conformational change of Ffh was also revealed to be important in the interaction between SecA and Ffh. However, MT had marginal effect on these conformational changes suggesting that the in vivo functionality of signal peptide is important in the interaction with Ffh and concomitant structural change of the protein.

• BMB Reports
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• v.39 no.5
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• pp.636-641
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• 2006

Our previous studies indicated that native carbonic anhydrase does not interact with hydrophobic adsorbents and that it acquires this ability upon denaturation. In the present study, an apo form of the enzyme was prepared by removal of zinc and a comparative study was performed on some characteristic features of the apo and native forms by far- and near-UV circular dichroism (CD), intrinsic fluorescent spectroscopy, 1-anilino naphthalene-8-sulfonate (ANS) binding, fluorescence quenching by acrylamide, and Tm measurement. Results indicate that protein flexibility is enhanced and the hydrophobic sites become more exposed upon conversion to the apo form. Accordingly, the apo structure showed a greater affinity for interaction with hydrophobic adsorbents as compared with the native structure. As observed for the native enzyme, heat denaturation of the apo form promoted interaction with alkyl residues present on the adsorbents and, by cooling followed by addition of zinc, catalytically-active immobilized preparations were obtained.

• Korean Journal of Pharmacognosy
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• v.33 no.4 s.131
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• pp.308-311
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• 2002

Advanced glycation end products(AGEs) are largly involved in the pathogenesis of diabetic nephropathy. It is obvious that inhibition of AGEs formation is important in preventing the occurrence and progression of diabetic nephropathy. In diabetes, this reaction is greatly accerated and is important in the pathogenesis of diabetic complications, especially diabetic nephropathy. Therefore, to seek possible AGEs inhibitors in herbal medicines, unprocessed - and processed Magnolia Bark were examined in vitro as basic data for aniaml experiment. The content of magnolol in unprocessed Magnolia Bark was $0.796{\pm}0.072%$, and after processing was decreased to $0.586{\pm}0.101%(p<0.01)$. The content of AGEs was measured by their intrinsic fluorescence. The $IC_{50}({\mu}g/ml)$ values of aminoguanidine, unprocessed- and procesled Magnolia Bark are $38.845{\pm}8.36{\mu}g/ml$, $54.264{\pm}3.153{\mu}g/ml$ and $27.882{\pm}1.836{\mu}g/ml$, respectively. This result means that prcessed Magnolia Bark was more effective than aminoguanidine, as positive control.

• BMB Reports
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• v.42 no.5
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• pp.293-298
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• 2009

Temperate mycobacteriophage L1 encodes an unusual repressor (CI) for regulating its lytic-lysogenic switching and, in contrast to the repressors of most temperate phages, it binds to multiple asymmetric operator DNAs. Here, ions like $Na^+$, $Cl^-$, and $acetate^-$ ions were demonstrated to facilitate the optimal binding of CI to cognate operator DNA, whereas $K^+$, $Li^+$, ${NH_4}^+$, $Mg^{2+}$, $carbonate^{2-}$, and $citrate^{3-}$ ions significantly affected its operator binding activity. Of these ions, $Mg^{2+}$ unfolded CI most severely at room temperature and, compared to $Mg^{2+}$, $Na^+$ provided improved thermal stability to CI. Furthermore, the intrinsic tryptophan fluorescence of CI was changed notably upon replacing $Na^+$ with $Mg^{2+}$ and these opposing effects of $Mg^{2+}$ and $Na^+$ were also noticed in their actions on the C-terminal fragment (CTD) of CI. Taken together, $Na^+$ appeared to be more appropriate than $Mg^{2+}$ for maintaining the biologically active conformation of CI needed for its optimal binding to operator DNA.