On the Difference in Action of the Laser Light with Wavelength Near 2mm on Biotissue in Gas and Water Media

It is shown that unlike action in the air environment, section of the biotissue in the water environment (physiological solution) is performed by the steam-gas stream which is formed as a result of superintensive boiling in thin (about 0.1 mm) a liquid layer in which absorbed laser radiation. Coagulation of the biotissue, adjacent to a section, happens due to heat which is produced via vapor condensation. Keywords: laser radiation in urology, a laser enucleation of the BPH, laser removal of the bladder cancer

Engineering of a wheat germ expression system to provide compatibility with a high throughput pET-based cloning platform

Wheat germ cell-free methods provide an important approach for the production of eukaryotic proteins. We have developed a protein expression vector for the TNT® SP6 High-Yield Wheat Germ Cell-Free (TNT WGCF) expression system (Promega) that is also compatible with our T7-based Escherichia coli intracellular expression vector pET15_NESG. This allows cloning of the same PCR product into either one of several pET_NESG vectors and this modified WGCF vector (pWGHisAmp) by In-Fusion LIC cloning (Zhu et al. in Biotechniques 43:354–359, 2007). Integration of these two vector systems allowed us to explore the efficacy of the TNT WGCF system by comparing the expression and solubility characteristics of 59 human protein constructs in both WGCF and pET15_NESG E. coli intracellular expression. While only 30% of these human proteins could be produced in soluble form using the pET15_NESG based system, some 70% could be produced in soluble form using the TNT WGCF system. This high success rate underscores the importance of eukaryotic expression host systems like the TNT WGCF system for eukaryotic protein production in a structural genomics sample production pipeline. To further demonstrate the value of this WGCF system in producing protein suitable for structural studies, we scaled up, purified, and analyzed by 2D NMR two 15N-, 13C-enriched human proteins. The results of this study indicate that the TNT WGCF system is a successful salvage pathway for producing samples of difficult-to-express small human proteins for NMR studies, providing an important complementary pathway for eukaryotic sample production in the NESG NMR structure production pipeline

Current challenges and future perspectives in oral absorption research: An opinion of the UNGAP network

Although oral drug delivery is the preferred administration route and has been used for centuries, modern drug discovery and development pipelines challenge conventional formulation approaches and highlight the insufficient mechanistic understanding of processes critical to oral drug absorption. This review presents the opinion of UNGAP scientists on four key themes across the oral absorption landscape: (1) specific patient populations, (2) regional differences in the gastrointestinal tract, (3) advanced formulations and (4) food-drug interactions. The differences of oral absorption in pediatric and geriatric populations, the specific issues in colonic absorption, the formulation approaches for poorly water-soluble (small molecules) and poorly permeable (peptides, RNA etc.) drugs, as well as the vast realm of food effects, are some of the topics discussed in detail. The identified controversies and gaps in the current understanding of gastrointestinal absorption-related processes are used to create a roadmap for the future of oral drug absorption research.European Cooperation in Science and Technology CA16205National Centre for Research and Development POIR.01.02.00-00-0011/17Ministry of Education, Youth and Sports of the Czech Republic TC19039, LTC18003Czech Science Foundation 18-00132

Инцидентальный рак предстательной железы

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Cell-free protein synthesis of membrane (1,3)-beta-D-glucan (curdlan) synthase: Co-translational insertion in liposomes and reconstitution in nanodiscs

A membrane-embedded curdlan synthase (CrdS) from Agrobacterium is believed to catalyse a repetitive addition of glucosyl residues from UDP-glucose to produce the (1,3)-β-d-glucan (curdlan) polymer. We report wheat germ cell-free protein synthesis (WG-CFPS) of full-length CrdS containing a 6xHis affinity tag and either Factor Xa or Tobacco Etch Virus proteolytic sites, using a variety of hydrophobic membrane-mimicking environments. Full-length CrdS was synthesised with no variations in primary structure, following analysis of tryptic fragments by MALDI-TOF/TOF Mass Spectrometry. Preparative scale WG-CFPS in dialysis mode with Brij-58 yielded CrdS in mg/ml quantities. Analysis of structural and functional properties of CrdS during protein synthesis showed that CrdS was co-translationally inserted in DMPC liposomes during WG-CFPS, and these liposomes could be purified in a single step by density gradient floatation. Incorporated CrdS exhibited a random orientation topology. Following affinity purification of CrdS, the protein was reconstituted in nanodiscs with Escherichia coli lipids or POPC and a membrane scaffold protein MSP1E3D1. CrdS nanodiscs were characterised by small-angle X-ray scattering using synchrotron radiation and the data obtained were consistent with insertion of CrdS into bilayers. We found CrdS synthesised in the presence of the Ac-AAAAAAD surfactant peptide or co-translationally inserted in liposomes made from E. coli lipids to be catalytically competent. Conversely, CrdS synthesised with only Brij-58 was inactive. Our findings pave the way for future structural studies of this industrially important catalytic membrane protein.Agalya Periasamy, Nadim Shadiac, Amritha Amalraj, Soňa Garajová, Yagnesh Nagarajan, Shane Waters, Haydyn D.T. Mertens, Maria Hrmov

The Center for Eukaryotic Structural Genomics

The Center for Eukaryotic Structural Genomics (CESG) is a “specialized” or “technology development” center supported by the Protein Structure Initiative (PSI). CESG’s mission is to develop improved methods for the high-throughput solution of structures from eukaryotic proteins, with a very strong weighting toward human proteins of biomedical relevance. During the first three years of PSI-2, CESG selected targets representing 601 proteins from Homo sapiens, 33 from mouse, 10 from rat, 139 from Galdieria sulphuraria, 35 from Arabidopsis thaliana, 96 from Cyanidioschyzon merolae, 80 from Plasmodium falciparum, 24 from yeast, and about 25 from other eukaryotes. Notably, 30% of all structures of human proteins solved by the PSI Centers were determined at CESG. Whereas eukaryotic proteins generally are considered to be much more challenging targets than prokaryotic proteins, the technology now in place at CESG yields success rates that are comparable to those of the large production centers that work primarily on prokaryotic proteins. We describe here the technological innovations that underlie CESG’s platforms for bioinformatics and laboratory information management, target selection, protein production, and structure determination by X-ray crystallography or NMR spectroscopy

Wheat germ cell-free expression system as a pathway to improve protein yield and solubility for the SSGCID pipeline

A set of 44 protein targets was used to test expression in the wheat germ cell-free system, the vast majority of which were expressed and soluble in this system; further increases in solubility were achieved by addition of the NVoy polymer