In vitro fluorescence studies of transcription factor IIB-DNA interaction

General transcription factor TFIIB is one of the basal constituents of the preinitiation complex of eukaryotic RNA polymerase II, acting as a bridge between the preinitiation complex and the polymerase, and binding promoter DNA in an asymmetric manner, thereby defining the direction of the transcription. Methods of fluorescence spectroscopy together with circular dichroism spectroscopy were used to observe conformational changes in the structure of recombinant human TFIIB after binding to specific DNA sequence. To facilitate the exploration of the structural changes, several site-directed mutations have been introduced altering the fluorescence properties of the protein. Our observations showed that binding of specific DNA sequences changed the protein structure and dynamics, and TFIIB may exist in two conformational states, which can be described by a different microenvironment of W52. Fluorescence studies using both intrinsic and exogenous fluorophores showed that these changes significantly depended on the recognition sequence and concerned various regions of the protein, including those interacting with other transcription factors and RNA polymerase II. DNA binding can cause rearrangements in regions of proteins interacting with the polymerase in a manner dependent on the recognized sequences, and therefore, influence the gene expression

Mechanism of polypurine tract primer generation by HIV-1 reverse transcriptase

HIV-1 reverse transcriptase (HIV-1 RT) possesses both DNA polymerase activity and RNase H activity that act in concert to convert single-stranded RNA of the viral genome to double-stranded DNA that is then integrated into the DNA of the infected cell. Reverse transcriptase-catalyzed reverse transcription critically relies on the proper generation of a polypurine tract (PPT) primer. However, the mechanism of PPT primer generation and the features of the PPT sequence that are critical for its recognition by HIV-1 RT remain unclear. Here, we used a chemical crosslinking method together with MD simulations and single-molecule assays to study the mechanism of PPT primer generation. We found that the PPT was specifically and properly recognized within covalently tethered HIV-1 RT-nucleic acid complexes. These findings indicated that recognition of the PPT occurs within a stable catalytic complex after its formation. We found that this unique recognition is based on two complementary elements that rely on the PPT sequence: RNase H sequence preference and incompatibility of the poly-rA/dT tract of the PPT with the nucleic acid conformation that is required for RNase H cleavage. The latter results from rigidity of the poly-rA/dT tract and leads to base-pair slippage of this sequence upon deformation into a catalytically relevant geometry. In summary, our results reveal an unexpected mechanism of PPT primer generation based on specific dynamic properties of the polyrA/dT segment and help advance our understanding of the mechanisms in viral RNA reverse transcription

1st International Electronic Conference on Food Science and Functional Foods

This research aimed to conduct a comparison of 8 different forms of brewery spent grain (BSG) on their polyphenolic content and antioxidant capacity as part of their potential as a functional food ingredient. The BSGs were dried until they reached a stable weight, grounded to pass through a 385 µm sieve and were vacuum packed in non-transparent packaging for further analysis. The results showed that BSG contained a high dietary fiber content which was dominated by a insoluble dietary fiber level of about 38.0–43.9% and a soluble dietary fiber content of about 3.9–9.6%. There were three groups of polyphenolic identified: flavan-3-ols, phenolic acids and flavonols at quantities of 362.1–1165.7 mg/kg, 65.8–122.5 mg/kg and 3.6–13.8 mg/kg, respectively. Antioxidant capacity was examined using an in vitro assessment: the 2,2′-Azinobis-(3-Ethylbenzthiazolin-6-Sulfonic Acid) (ABTS) capacity ranged from 0.086 to 0.241 mmol Trolox/100 g while the ferric reducing antioxidant potential (FRAP) capacity ranged from 0.106 to 0.306 µmol TE/100 g. In conclusion, BSG as a brewery waste can potentially be used as a functional food ingredient due to its properties. It is suggested that further studies are needed to explore BSG’s impact on the development of functional food products.Keywords: brewery spent grain; valorization; agricultural by-products; functional food; dietary fiber; polyphenolic compounds; antioxidant</p

Ewolucja rozwoju systemu pomocy społecznej w ujęciu ekonomicznym na obszarze województwa opolskiego w latach 2008-2016

The aim of the article is to approximate the changes that took place in the field of social assistance in 2008-2016 in the Opolskie Voivodeship. The purpose of the application is to show the economic consequences of the evolution of the social assistance system in the Opolskie Voivodeship in 2008-2016. The research method used in the work covers both the analysis of the literature on the subject and the analysis of statistical data. The conducted research allows to conclude that the limitation of unfavorable phenomena such as unemployment and poverty influenced the change of the social welfare beneficiary profile - from an unemployed person to a disabled, seriously ill or elderly person. The change of the character of social assistance is connected with the necessity of reorganization of the support system in the voivodship towards the development of social services, above all caring, nursing or supportive services, at the same time retaining both monetary and non-monetary financial support.(original abstract

Evolution of the development of the social welfare system in economic consideration in the Opolskie Voivodeship in the years 2008-2016

Celem artykułu jest przybliżenie zmian, jakie dokonały się na gruncie pomocy społecznej w latach 2008-2016 na terenie województwa opolskiego. Celem aplikacyjnym jest ukazanie ekonomicznych skutków ewolucji systemu pomocy społecznej na terenie województwa opolskiego w latach 2008-2016. Wykorzystana w pracy metoda badawcza obejmuje zarówno analizę literatury tematu, jak i analizę danych statystycznych. Wykonane badania pozwalają stwierdzić, że ograniczenie niekorzystnych zjawisk, takich jak bezrobocie oraz ubóstwo, wpłynęło na zmianę profilu beneficjenta pomocy społecznej – od osoby bezrobotnej, do osoby niepełnosprawnej, ciężko chorej czy też starszej. Zmiana charakteru pomocy społecznej wiąże się z koniecznością reorganizacji systemu wsparcia na terenie województwa w kierunku rozwoju usług społecznych, przede wszystkim opiekuńczych, pielęgnacyjnych czy też wspomagających, przy równoczesnym zachowaniu pomocy materialnej, zarówno pieniężnej, jak i niepieniężnej.The aim of the article is to approximate the changes that took place in the field of social assistance in 2008-2016 in the Opolskie Voivodeship. The purpose of the application is to show the economic consequences of the evolution of the social assistance system in the Opolskie Voivodeship in 2008-2016. The research method used in the work covers both the analysis of the literature on the subject and the analysis of statistical data. The conducted research allows to conclude that the limitation of unfavorable phenomena such as unemployment and poverty influenced the change of the social welfare beneficiary profile – from an unemployed person to a disabled, seriously ill or elderly person. The change of the character of social assistance is connected with the necessity of reorganization of the support system in the voivodship towards the development of social services, above all caring, nursing or supportive services, at the same time retaining both monetary and non-monetary financial support

Coordination between the polymerase and RNase H activity of HIV-1 reverse transcriptase

Replication of human immunodeficiency virus 1 (HIV-1) involves conversion of its single-stranded RNA genome to double-stranded DNA, which is integrated into the genome of the host. This conversion is catalyzed by reverse transcriptase (RT), which possesses DNA polymerase and RNase H domains. The available crystal structures suggest that at any given time the RNA/DNA substrate interacts with only one active site of the two domains of HIV-1 RT. Unknown is whether a simultaneous interaction of the substrate with polymerase and RNase H active sites is possible. Therefore, the mechanism of the coordination of the two activities is not fully understood. We performed molecular dynamics simulations to obtain a conformation of the complex in which the unwound RNA/DNA substrate simultaneously interacts with the polymerase and RNase H active sites. When the RNA/DNA hybrid was immobilized at the polymerase active site, RNase H cleavage occurred, experimentally verifying that the substrate can simultaneously interact with both active sites. These findings demonstrate the existence of a transient conformation of the HIV-1 RT substrate complex, which is important for modulating and coordinating the enzymatic activities of HIV-1 RT