Whole-cell fungal transformation of precursors into dyes

<p>Abstract</p> <p>Background</p> <p>Chemical methods of producing dyes involve extreme temperatures and unsafe toxic compounds. Application of oxidizing enzymes obtained from fungal species, for example laccase, is an alternative to chemical synthesis of dyes. Laccase can be replaced by fungal biomass acting as a whole-cell biocatalyst with properties comparable to the isolated form of the enzyme. The application of the whole-cell system simplifies the transformation process and reduces the time required for its completion. In the present work, four fungal strains with a well-known ability to produce laccase were tested for oxidation of 17 phenolic and non-phenolic precursors into stable and non-toxic dyes.</p> <p>Results</p> <p>An agar-plate screening test of the organic precursors was carried out using four fungal strains: <it>Trametes versicolor</it>, <it>Fomes fomentarius</it>, <it>Abortiporus biennis</it>, and <it>Cerrena unicolor</it>. Out of 17 precursors, nine were transformed into coloured substances in the presence of actively growing fungal mycelium. The immobilized fungal biomass catalyzed the transformation of 1 mM benzene and naphthalene derivatives in liquid cultures yielding stable and non-toxic products with good dyeing properties. The type of fungal strain had a large influence on the absorbance of the coloured products obtained after 48-hour transformation of the selected precursors, and the most effective was <it>Fomes fomentarius </it>(<it>FF25</it>). Whole-cell transformation of AHBS (3-amino-4-hydroxybenzenesulfonic acid) into a phenoxazinone dye was carried out in four different systems: in aqueous media comprising low amounts of carbon and nitrogen source, in buffer, and in distilled water.</p> <p>Conclusions</p> <p>This study demonstrated the ability of four fungal strains belonging to the ecological type of white rot fungi to transform precursors into dyes. This paper highlights the potential of fungal biomass for replacing isolated enzymes as a cheaper industrial-grade biocatalyst for the synthesis of dyes and other commercially important products. The use of immobilized fungal biomass limits free migration of cells and facilitates their reuse in a continuous system for precursor transformation.</p

Mapping the Human Brain in Frequency Band Analysis of Brain Cortex Electroencephalographic Activity for Selected Psychiatric Disorders

There are still no good quantitative methods to be applied in psychiatric diagnosis. The interview is still the main and most important tool in the psychiatrist work. This paper presents the results of electroencephalographic research with the subjects of a group of 30 patients with psychiatric disorders compared to the control group of healthy volunteers. All subjects were solving working memory task. The digit-span working memory task test was chosen as one of the most popular tasks given to subjects with cognitive dysfunctions, especially for the patients with panic disorders, depression (including the depressive phase of bipolar disorder), phobias, and schizophrenia. Having such cohort of patients some results for the subjects with insomnia and Asperger syndrome are also presented. The cortical activity of their brains was registered by the dense array EEG amplifier. Source localization using the photogrammetry station and the sLORETA algorithm was then performed in five EEG frequency bands. The most active Brodmann Areas are indicated. Methodology for mapping the brain and research protocol are presented. The first results indicate that the presented technique can be useful in finding psychiatric disorder neurophysiological biomarkers. The first attempts were made to associate hyperactivity of selected Brodmann Areas with particular disorders

SYK inhibition targets acute myeloid leukemia stem cells by blocking their oxidative metabolism

Spleen tyrosine kinase (SYK) is an important oncogene and signaling mediator activated by cell surface receptors crucial for acute myeloid leukemia (AML) maintenance and progression. Genetic or pharmacologic inhibition of SYK in AML cells leads to increased differentiation, reduced proliferation, and cellular apoptosis. Herein, we addressed the consequences of SYK inhibition to leukemia stem-cell (LSC) function and assessed SYK-associated pathways in AML cell biology. Using gain-of-function MEK kinase mutant and constitutively active STAT5A, we demonstrate that R406, the active metabolite of a small-molecule SYK inhibitor fostamatinib, induces differentiation and blocks clonogenic potential of AML cells through the MEK/ERK1/2 pathway and STAT5A transcription factor, respectively. Pharmacological inhibition of SYK with R406 reduced LSC compartment defined as CD34+CD38-CD123+ and CD34+CD38-CD25+ in vitro, and decreased viability of LSCs identified by a low abundance of reactive oxygen species. Primary leukemic blasts treated ex vivo with R406 exhibited lower engraftment potential when xenotransplanted to immunodeficient NSG/J mice. Mechanistically, these effects are mediated by disturbed mitochondrial biogenesis and suppression of oxidative metabolism (OXPHOS) in LSCs. These mechanisms appear to be partially dependent on inhibition of STAT5 and its target gene MYC, a well-defined inducer of mitochondrial biogenesis. In addition, inhibition of SYK increases the sensitivity of LSCs to cytarabine (AraC), a standard of AML induction therapy. Taken together, our findings indicate that SYK fosters OXPHOS and participates in metabolic reprogramming of AML LSCs in a mechanism that at least partially involves STAT5, and that SYK inhibition targets LSCs in AML. Since active SYK is expressed in a majority of AML patients and confers inferior prognosis, the combination of SYK inhibitors with standard chemotherapeutics such as AraC constitutes a new therapeutic modality that should be evaluated in future clinical trials

Role of Donor Activating KIR–HLA Ligand–Mediated NK Cell Education Status in Control of Malignancy in Hematopoietic Cell Transplant Recipients

AbstractSome cancers treated with allogeneic hematopoietic stem cell transplantation (HSCT) are sensitive to natural killer cell (NK) reactivity. NK function depends on activating and inhibitory receptors and is modified by NK education/licensing effect and mediated by coexpression of inhibitory killer-cell immunoglobulin-like receptor (KIR) and its corresponding HLA I ligand. We assessed activating KIR (aKIR)-based HLA I–dependent education capacity in donor NKs in 285 patients with hematological malignancies after HSCT from unrelated donors. We found significantly adverse progression-free survival (PFS) and time to progression (TTP) in patients who received transplant from donors with NKs educated by C1:KIR2DS2/3, C2:KIR2DS1, or Bw4:KIR3DS1 pairs (for PFS: hazard ratio [HR], 1.70; P = .0020, Pcorr = .0039; HR, 1.54; P = .020, Pcorr = .039; HR, 1.51; P = .020, Pcorr = .040; and for TTP: HR, 1.82; P = .049, Pcorr = .096; HR, 1.72; P = .096, Pcorr = .18; and HR, 1.65; P = .11, Pcorr = .20, respectively). Reduced PFS and TTP were significantly dependent on the number of aKIR-based education systems in donors (HR, 1.36; P = .00031, Pcorr = .00062; and HR, 1.43; P = .019, Pcorr = .038). Furthermore, the PFS and TTP were strongly adverse in patients with missing HLA ligand cognate with educating aKIR-HLA pair in donor (HR, 3.25; P = .00022, Pcorr = .00045; and HR, 3.82; P = .027, Pcorr = .054). Together, these data suggest important qualitative and quantitative role of donor NK education via aKIR-cognate HLA ligand pairs in the outcome of HSCT. Avoiding the selection of transplant donors with high numbers of aKIR-HLA-based education systems, especially for recipients with missing cognate ligand, is advisable

Bioactive Properties of a Novel Antibacterial Dye Obtained from Laccase-Mediated Oxidation of 8-Anilino-1-naphthalenesulfonic Acid

Fungal laccase obtained from a Cerrena unicolor strain was used as an effective biocatalyst for the transformation of 8-anilino-1-naphthalenesulfonic acid into a green-coloured antibacterial compound, which can be considered as both an antimicrobial agent and a textile dye, simultaneously. The process of biosynthesis was performed in buffered solutions containing methanol as a co-solvent, allowing better solubilisation of substrate. The transformation process was optimised in terms of the buffer pH value, laccase activity, and concentrations of the substrate and co-solvent. The crude product obtained exhibited low cytotoxicity, antibacterial properties against Staphylococcus aureus and Staphylococcus epidermidis, and antioxidant properties. Moreover, the synthesised green-coloured compound proved non-allergenic and demonstrated a high efficiency of dyeing wool fibres

Wpływ procesu dojrzewania i zawartości tłuszczu na zmiany poziomu witaminy k w serach®

In this study, the vitamin K1 (phylloquinone) and vitamin K2 (menaquinone) contents of a selected range of cheeses were measured by high-performance liquid chromatography (HPLC) and compared with the fat content, maturation time and origin of the cheeses. In our study, the highest vitamin K2 content was recorded in Gouda (678.12 ng/g), Edam (712.70 ng/g) and Emmentaler (733.10 ng/g) cheeses with comparable levels of vitamin K1 in the analysed products (31.60 ng/g, 34.63 ng/g and 24.35 ng/g, respectively) and fat content (27%, 28% and 30%, respectively), as well as in Gouda cheese with a fat content of 48% after 48 weeks of maturation (756.50 ng/g). The fat content of the cheese was a factor that influenced the vitamin K content, with products with lower fat content having lower total vitamin K content compared to the other products analysed. Given the reports that poor vitamin K status is one of the risk factors for cardiovascular disease in the absence of conclusive evidence of adverse cardiovascular effects of dairy fats, cheese should be considered as an important dietary component for those concerned about heart health.W tym badaniu oznaczono zawartość witaminy K1 (filochinonu) i witaminy K2 (menachinonu) w wybranej gamie serów metodą wysokosprawnej chromatografii cieczowej (HPLC) i porównano z zawartością tłuszczu, czasem dojrzewania i pochodzeniem serów. W badaniach własnych największą zawartość witaminy K2 odnotowano w serach Gouda (678,12 ng/g), Edam (712,70 ng/g) i Emmentaler (733,10 ng/g) przy porównywalnych poziomach witaminy K1 w analizowanych produktach (odpowiednio 31,60 ng/g, 34,63 ng/g i 24,35 ng/g) i zawartości tłuszczu (odpowiednio 27%, 28% i 30%), jak również w serze Gouda o zawartości tłuszczu 48% po 48 tygodniach dojrzewania (756.50 ng/g). Zawartość tłuszczu w serze była czynnikiem, który wpływał na zawartość witaminy K, przy czym produkty o niższej zawartości tłuszczu miały niższą całkowitą zawartość witaminy K w porównaniu z pozostałymi analizowanymi produktami. Biorąc pod uwagę doniesienia, że słaby status witaminy K jest jednym z czynników ryzyka chorób sercowo-naczyniowych przy braku jednoznacznych dowodów na niekorzystny wpływ tłuszczów mlecznych na układ krążenia, sery powinny być traktowane jako ważny składnik diety osób, które dbają o zdrowie serca

Genetic Characterization of the CcpA-Dependent, Cellobiose-Specific PTS System Comprising CelB, PtcB and PtcA that Transports Lactose in Lactococcus lactis IL1403.

Lactose metabolism is one of the most important areas of research on Lactic Acid Bacteria (LAB). In rapidly acidifying industrial Lactococcus lactis strains, lactose is transported by a lactose-specific phosphotransferase system (PTS) encoded by a plasmid. However, an alternative lactose catabolic pathway was evidenced in the plasmid-cured, and thus initially lactose-negative L. lactis IL1403. We showed that in this strain the chromosomally-encoded cellobiose-specific PTS system comprising the celB, ptcB and ptcA genes is also able to transport lactose. By expression studies in the wild type IL1403 strain and IBB550, its ccpA-deficient derivative, we demonstrated that celB, ptcB and ptcA are tightly regulated by the general catabolite repression system, whereas celB additionally requires the presence of cellobiose to be fully induced. The comparison of expression levels of sugar catabolic genes indicated that the efficiency of CcpA-mediated catabolic repression depends on conservation of the cre sequence, and that in the case of perfect matching with the cre consensus, CcpA still drives a strong repression even under non-repressing conditions

Genetic Characterization of the CcpA-Dependent, Cellobiose-Specific PTS System Comprising CelB, PtcB and PtcA that Transports Lactose in Lactococcus lactis IL1403.

Lactose metabolism is one of the most important areas of research on Lactic Acid Bacteria (LAB). In rapidly acidifying industrial Lactococcus lactis strains, lactose is transported by a lactose-specific phosphotransferase system (PTS) encoded by a plasmid. However, an alternative lactose catabolic pathway was evidenced in the plasmid-cured, and thus initially lactose-negative L. lactis IL1403. We showed that in this strain the chromosomally-encoded cellobiose-specific PTS system comprising the celB, ptcB and ptcA genes is also able to transport lactose. By expression studies in the wild type IL1403 strain and IBB550, its ccpA-deficient derivative, we demonstrated that celB, ptcB and ptcA are tightly regulated by the general catabolite repression system, whereas celB additionally requires the presence of cellobiose to be fully induced. The comparison of expression levels of sugar catabolic genes indicated that the efficiency of CcpA-mediated catabolic repression depends on conservation of the cre sequence, and that in the case of perfect matching with the cre consensus, CcpA still drives a strong repression even under non-repressing conditions

Influence of Carrier Structure and Physicochemical Factors on Immobilisation of Fungal Laccase in Terms of Bisphenol A Removal

Laccase from Pleurotus ostreatus was immobilised on porous Purolite&reg; carriers and amino-functionalised ultrafiltration membranes. The results indicated a correlation between the carrier structure and the activity of laccase immobilised thereon. The highest activity was obtained for carriers characterised by a small particle size and a larger pore diameter (the porous carriers with an additional spacer (C2 and C6) and octadecyl methacrylate beads with immobilised laccase activity of 5.34 U/g, 2.12 U/g and 7.43 U/g, respectively. The conditions of immobilisation and storage of immobilised laccase were modified to improve laccase activity in terms of bisphenol A transformation. The highest laccase immobilisation activity was obtained on small bead carriers with a large diameter of pores incubated in 0.1 M phosphate buffer pH 7 and for immobilisation time of 3 h at 22 &deg;C. The immobilised LAC was stable for four weeks maintaining 80&ndash;90% of its initial activity in the case of the best C2, C6, and C18 carriers. The immobilised laccase transformed 10 mg/L of BPA in 45% efficiency and decreased its toxicity 3-fold in the Microtox tests. The effectiveness of BPA transformation, and the legitimacy of conducting this process due to the reduction of the toxicity of the resulting reaction products have been demonstrated. Reusability of immobilised LAC has been proven during BPA removal in 10 subsequent batches