Purification,characterization, and identification of a novel bifunctional catalase-phenol oxidase from <Emphasis Type="Italic">Scytalidium thermophilum</Emphasis>

A novel bifunctional catalase with an additional phenol oxidase activity was isolated from a thermophilic fungus, Scytalidium thermophilum. This extracellular enzyme was purified ca. 10-fold with 46% yield and was biochemically characterized. The enzyme contains heme and has a molecular weight of 320 kDa with four 80 kDa subunits and an isoelectric point of 5.0. Catalase and phenol oxidase activities were most stable at pH 7.0. The activation energies of catalase and phenol oxidase activities of the enzyme were found to be 2.7 +/- 0.2 and 10.1 +/- 0.4 kcal/mol, respectively. The pure enzyme can oxidize o-diphenols such as catechol, caffeic acid, and L-DOPA in the absence of hydrogen peroxide and the highest oxidase activity is observed against catechol. No activity is detected against tyrosine and common laccase substrates such as ABTS and syringaldazine with the exception of weak activity with p-hydroquinone. Common catechol oxidase inhibitors, salicylhydroxamic acid and p-coumaric acid, inhibit the oxidase activity. Catechol oxidation activity was also detected in three other catalases tested, from Aspergillus niger, human erythrocyte, and bovine liver, suggesting that this dual catalase-phenol oxidase activity may be a common feature of catalases.     

Protective role of carnitine in breast cancer via decreasing arginase activity and increasing nitric oxide

Breast cancer remains one of the most common types of cancer. High levels of arginase and ornithine in different carcinomas may indicate their relation to cancer. Carnitine is a cofactor required for the transformation of free long-chain fatty acids into acetyl-carnitines. We have examined the protective effect of carnitine and the possibility that it disturbs arginase-nitric oxide (NO) interaction. Histopathological examination, arginase activity, ornithine and NO levels were determined in tumour tissues. Mitotic cells significantly decreased in the treatment group. Tissue arginase activity and ornithine levels decreased significantly with carnitine. NO levels were significantly higher in the treatment group. One of the possible mechanisms of carnitine's protective role in tumour progression might be its promotion of NO. This mechanism could decrease the production of tumour-promoting agents, polyamines, and increase the production of NO, thereby exerting a protective effect on cancer development.     

Differential proteome and cellular adhesion analyses of the probiotic bacterium Lactobacillus acidophilus NCFM grown on raffinose – an emerging prebiotic

Whole cell and surface proteomes were analyzed together with adhesive properties of the probiotic bacterium Lactobacillus acidophilus NCFM (NCFM) grown on the emerging prebiotic raffinose, exemplifying a synbiotic. Adhesion of NCFM to mucin and intestinal HT‐29 cells increased three‐fold after culture with raffinose versus glucose, as also visualized by scanning electron microscopy. Comparative proteomics using 2D‐DIGE showed 43 unique proteins to change in relative abundance in whole cell lysates from NCFM grown on raffinose compared to glucose. Furthermore, 14 unique proteins in 18 spots of the surface subproteome underwent changes identified by differential 2DE, including elongation factor G, thermostable pullulanase, and phosphate starvation inducible stress‐related protein increasing in a range of +2.1 ? +4.7 fold. By contrast five known moonlighting proteins decreased in relative abundance by up to ?2.4 fold. Enzymes involved in raffinose catabolism were elevated in the whole cell proteome; α‐galactosidase (+13.9 fold); sucrose phosphorylase (+5.4 fold) together with metabolic enzymes from the Leloir pathway for galactose utilization and the glycolysis; β‐galactosidase (+5.7 fold); galactose (+2.9/+3.1 fold) and fructose (+2.8 fold) kinases. The insights at the molecular and cellular levels contributed to the understanding of the interplay of a synbiotic composed of NCFM and raffinose with the host.     

Investigation of VHL gene associated with miR-223 in clear cell renal cell carcinoma

Background

Clear cell type renal cell carcinoma (ccRCC) is the most common renal cell carcinoma (RCC). In this study, we examined the expressions of VHL and miR-223 in ccRCC patients? tissues to investigate the possible role in the development of ccRCC.

Methods and results

This study collected five expression profiles (GSE36139, GSE3, GSE73731, GSE40435, and GSE26032) from Gene Omnibus Data. Expressions of VHL and miR-223 in paraffinized tumor and normal tissues of 100 Turkish patients' ccRCC tissues were determined by bioinformatic data mining and real-time quantitative polymerase chain reaction (qRT-PCR). The VHL gene was subjected to mutational analysis by DNA sequencing, and pVHL was analyzed using western blotting. Our study's t-test and Pearson correlation analysis showed that VHL gene expression in tumoral tissues with a???0.39-fold decrease was not significantly lower than normal tissues (p?=?0.441), and a 0.97-fold increase miR-223 (p?=?0.045) was determined by real-time PCR. Also, as a result of DNA sequence analysis performed in the VHL gene, it was found that 26% of the patients have mutations. The mutations for (VHL):c.60C>A (p.Val20=) and (VHL):c.467delA (p.Tyr156Leu) was detected for the first time in Turkish patients.

Conclusions

The present study demonstrated that the differences in the expression levels of miR-223 have the potential to be biomarkers to determine the poor prognosis in ccRCC.

     

Effect of cobalt-60 (γ radiation) on multidrug-resistant multiple myeloma cell lines

Emergence of resistance to chemotherapy and radiotherapy is a major obstacle for the successful treatment of MM (multiple myeloma). Prednisone, vincristine and melphalan are commonly used chemotherapeutic agents for the treatment of MM. In the current study, we examined the presence of possible cross-resistance between these drugs and gamma (γ) radiation. Prednisone, vincristine and melphalan resistant RPMI-8226 and U-266 MM cells were generated by stepwise increasing concentrations of the drugs. The sensitive and resistant cells were exposed to 200- and 800 cGy γ radiation, and proliferation was examined by XTT {2,3-bis(2-methoxy-4-nitro-5-sulfophenyl)-5-[(phenylamino)carbonyl]-2H-tetrazolium hydroxide} assay. The results showed that Prednisone- and melphalan-resistant RPMI-8226 cells were also cross-resistant to 200 and 800 cGy γ radiation application, while vincristine-resistant cells did not show resistance. On the other hand, Prednisone-, vincristine- and melphalan-resistant U-266 cells showed cross-resistance to 200- and 800 cGy γ radiation application. These results demonstrated that MM cells resistant to anticancer agents respond to radiation in different levels. These findings may be important in the clinical applications of radiation therapy in the treatment of vincristine resistant MM.     

Effects of the ecological conditions on morphological variations of the green toad, Bufo viridis, in Turkey

Ten populations of the green toad, Bufo viridis, from three areas having different climatic regimes were compared. Multivariate analyses of morphological characteristics revealed three distinct populations of green toads: the first group inhabiting the lowland and humid localities was characterized by a small body size, the second group occurred at higher altitudes and in desert areas had a significantly larger body size, while the third group from the Mediterranean localities was characterized by a large tympanum, a longer tarsus and short first toes. Interestingly, the specimens from Giresun Island were clustered with the second group with a similar pattern of body size. Differences in the morphological characteristics among populations were positively associated with ecological conditions, and not with geographic distance. Some characteristics previously used for taxonomic purposes in B. viridis were shown to be associated with local climatic conditions.