Application of Ni(II)-Assisted Peptide Bond Hydrolysis to Non-Enzymatic Affinity Tag Removal

In this study, we demonstrate a non-enzymatic method for hydrolytic peptide bond cleavage, applied to the removal of an affinity tag from a recombinant fusion protein, SPI2-SRHWAP-His6. This method is based on a highly specific Ni(II) reaction with (S/T)XHZ peptide sequences. It can be applied for the protein attached to an affinity column or to the unbound protein in solution. We studied the effect of pH, temperature and Ni(II) concentration on the efficacy of cleavage and developed an analytical protocol, which provides active protein with a 90% yield and ∼100% purity. The method works well in the presence of non-ionic detergents, DTT and GuHCl, therefore providing a viable alternative for currently used techniques

Interaction patterns of methoprene-tolerant and germ cell-expressed Drosophila JH receptors suggest significant differences in their functioning

Methoprene-tolerant (Met) and germ cell-expressed (Gce) proteins were shown to be juvenile hormone (JH) receptors of Drosophila melanogaster with partially redundant functions. We raised the question of where the functional differentiation of paralogs comes from. Therefore, we tested Met and Gce interaction patterns with selected partners. In this study, we showed the ability of Gce and its C-terminus (GceC) to interact with 14-3-3 in the absence of JH. In contrast, Met or Met C-terminus (MetC) interactions with 14-3-3 were not observed. We also performed a detailed structural analysis of Met/Gce interactions with the nuclear receptor fushi tarazu factor-1 (Ftz-F1) ligand-binding domain. We showed that GceC comprising an Ftz-F1-binding site and full-length protein interacts with Ftz-F1. In contrast to Gce, only MetC (not full-length Met) can interact with Ftz-F1 in the absence of JH. We propose that the described differences result from the distinct tertiary structure and accessibility of binding sites in the full-length Met/Gce. Moreover, we hypothesize that each interacting partner can force disordered MetC and GceC to change the structure in a partner-specific manner. The observed interactions seem to determine the subcellular localization of Met/Gce by forcing their translocation between the nucleus and the cytoplasm, which may affect the activity of the proteins. The presented differences between Met and Gce can be crucial for their functional differentiation during D. melanogaster development and indicate Gce as a more universal and more active paralog. It is consistent with the theory indicating gce as an ancestor gene

Use of satellite data in monitoring of hydrophysical parameters of the baltic sea environment

Intensive development of infrastructure for fast processing of outsized amount of space-borne data enables now to use the satellite data for operational controlling the state of its environment. In our presentation we show some examples of analysis of processes in marine environment which are possible due to satellite data and algorithms of its processing developed in SatBaltic Project. It concerns supporting of modelling of solar energy inflow to the sea with space-borne input data, identification and analysis of sea ice cover, supporting of oil spill detection, and identification of phenomena which modify spatial distribution of the sea surface temperature

Daily radiation budget of the Baltic Sea surface from satellite data

Recently developed system for assessment of radiation budget for the Baltic Sea has been presented and verified. The system utilizes data from various sources: satellite, model and in situ measurements. It has been developed within the SatBałtyk project (Satellite Monitoring of the Baltic Sea Environment - www.satbaltyk.eu) where the energy radiation budget is one of the key element. The SatBałtyk system generates daily maps of the all components of radiation budget on every day basis. We show the scheme of making daily maps, applied algorithms and empirical data collection within the system. An empirical verification of the system has been carried out based on empirical data collected on the oil rig placed on the Baltic Sea. This verification concerned all the components of the surface radiation budget. The average daily NET products are estimated with statistical error ca. 13 Wm-2. The biggest absolute statistical error is for LWd component and equals 14 Wm-2. The relative error in relation to the average annual values for whole Baltic is the biggest for SWu and reaches 25%. All estimated components have correlation coefficient above 0.91

Practical applicability and preliminary results of the Baltic environmental satellite remote sensing system (SATBAŁTYK)

The SatBałtyk (Satellite Monitoring of the Baltic Sea Environment) project is being realized in Poland by the SatBałtyk Scientific Consortium, specifically appointed for this purpose, which associates four scientific institutions: the Institute of Oceanology PAN in Sopot – coordinator of the project, the University of Gdańsk (Institute of Oceanography), the Pomeranian Academy in Słupsk (Institute of Physics) and the University of Szczecin (Institute of Marine Sciences). The project is aiming to prepare a technical infrastructure and set in motion operational procedures for the satellite monitoring of the Baltic Sea ecosystem. The main sources of input data for this system will be the results of systematic observations by metrological and environmental satellites such as TIROS N/NOAA, MSG (currently Meteosat 10), EOS/AQUA and Sentinel -1, 2, 3 (in the future). The system will deliver on a routine basis the variety of structural and functional properties of this sea, based on data provided by relevant satellites and supported by hydro-biological models. Among them: the solar radiation influx to the sea’s waters in various spectral intervals, energy balances of the short- and long-wave radiation at the Baltic Sea surface and in the upper layers of the atmosphere over the Baltic, sea surface temperature distribution, dynamic states of the water surface, concentrations of chlorophyll a and other phytoplankton pigments in the Baltic waters, spatial distributions of algal blooms, the occurrence of coastal upwelling events, and the characteristics of primary production of organic matter and photosynthetically released oxygen in the water and many others. The structure of the system and preliminary results will be presented

Coordination properties of dithiobutylamine (DTBA), a newly introduced protein disulfide reducing agent

The acid-base properties and metal-binding abilities of (2S)-2-amino-1,4-dimercaptobutane, otherwise termed dithiobutylamine (DTBA), which is a newly introduced reagent useful for reducing protein and peptide disulfides, were studied in solution using potentiometry, (1)H NMR spectroscopy, spectropolarimetry, and UV-vis spectroscopy. The list of metal ions studied here includes Zn(II), Cd(II), Ni(II), Co(II), and Cu(I). We found that DTBA forms specific and very stable polynuclear and mononuclear complexes with all of these metal ions using both of its sulfur donors. DTBA forms complexes more stable than those of the commonly used disulfide reducing agent DTT, giving it more interference capacity in studies of metal binding in thiol-containing biomolecules. The ability of DTBA to strongly bind metal ions is reflected in its limited properties as a thiol protectant in their presence, which is manifested through slower disulfide reduction kinetics. We found that this effect correlated with the stabilities of the complexes. Additionally, the reducing properties of DTBA toward MMTS-modified papain (MMTS = S-methylmethanethiosulfonate) were also significantly affected by the investigated metal ions. In this case, however, electrostatic interactions and stereospecific effects, rather than metal-binding abilities, were found to be responsible for the reduced protective properties of DTBA. Despite its limitations, a high affinity toward metal ions makes DTBA an attractive agent in competition studies with metalloproteins

Effect of the temperament of dairy cows on lifetime production efficiency

The aim of this study was to analyse the effect of the temperament of cows on their lifetime production efficiency. Data on milk performance, longevity and culling reasons of 11 629 Polish Holstein-Friesian cows were extracted from the SYMLEK database. Using the methodology of the Polish Federation of Cattle Breeders and Dairy Farmers, cow temperament was assessed as follows: 1 – calm; 2 – normal; and 3 – excitable or aggressive. Most cows showed a normal temperament (89.52 %). Over successive years of the evaluation, there were increases in the proportion of cows with a calm (from 2.98 to 6.85 %) and excitable temperament (from 7.50 to 8.27 %). Temperament was found to have a highly significant effect on first lactation milk yield, first lactation daily milk yield, milk yield per day of age, and milk yield per day of productive life. In cows with a calm temperament, first lactation milk yield was 621 kg higher than in cows with a normal temperament and 329 kg higher compared to excitable (aggressive) cows