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Microsatellite region in the murine oncogene src - its primary structure and variability during evolution
Fučík, Vladimír ; Šindelka, Radek ; Beran, Jaroslav ; Ferjentsik, Zoltán ; Jonák, Jiří
In the fifth intron of the Mus musculus oncogene src a complex microsatellite was discovered that consists of a CA run and 31 bp apart followed by track of TG dinucleotides interrupted by extra Gs in specific intervals. As the 31 bp interlink can assume a hairpin structure, the whole region can be viewed as an imperfect inverted repeat. The microsatellite was sequenced in imbred lines of M. m. musculus and M. m. domesticus, as well as in mice of both subspecies trapped near the hybrid zone. Differences in the length of repeat regions, insertion/deletions and simple base changes characterize the strains. Hybrids between the two species could be identified by sequencing of cloned PCR products. The structure of the microsatellite in more distant species was found to be conspicuously different. Instead of frequent (CA)18-25, we can see only (CA)8 in M. spretus, CATGCGCCCCCCA in Mus pahari and CACCCCCC in Mus caroli. No similar structure is present in the corresponding human src intron.
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Experimental methods for phase determination /MIR,MAD,SAD/ in crystallography of biological macromolecules
Brynda, Jiří
Importance of unbiased phases in post genomic era is emphasized. Article describes the classical methods of phase determination: 1) Multiple Isomorphous replacement, heavy-atom derivative (MIR) 2) Multiple Anomalous Dispersion (MAD) Phasing 3) Single Anomalous Dispersion (SAD) Phasing The better your initial, experimental map is, the more model you will be able to build confidently into this unbiased map before proceeding to phase combination. MAD phases result from perfectly isomorphous data, and do not deteriorate at low resolution as do MIR phases. MIR phases, on the other hand, provide superior phasing power at low resolution resulting in better connectivity. Not surprising, the best maps are either MAD or combined MIR-MAD maps.
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Processing of Diffraction Data Obtained by Oscillation Method on Crystals of Biological Macromolecules
Brynda, Jiří
The components of monochromatic diffractometer suitable for data collection on crystals of biological macromolecules are described, as well as basics of rotation method. The optimal strategy for collecting X-ray diffraction data from macromolecular crystals is discussed. The geometric factors influencing the completeness of data are arising from the symmetry of the reciprocal lattice and from the experimental setup; they affect quantitatively the completeness of the measured set of reflections.
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A Phenylnorstatine Inhibitor Binding to HIV-1 Protease: Geometry, Protonation and Subsite-Pocket Interactions Analyzed at Atomic Resolution
Brynda, Jiří ; Řezáčová, Pavlína ; Fábry, Milan ; Hořejší, Magdalena ; Štouračová, Renata ; Sedláček, Juraj ; Souček, M. ; Hradílek, M. ; Lepšík, M. ; Konvalinka, J.
The x-ray structure of a complex of HIV-1 protease (PR) with a phenylnorstatine inhibitor Z-Pns-Phe-Glu-Glu-NH2 has been determined at 1.03 A, the highest resolution so far reported for any HIV PR complex. The inhibiot shows subnanomolar Ki values for both the wild-type PR and the variant representing one of the most common mutations linked to resistance developoment. The structure displays a unique pattern of hydrogen bonding to the two catalytic aspartate residues. The high resolution permits to assess the donor/acceptor realtions of this hydrogen bonding and to indicate a proton shared by the two catalytic residues. Structural mechanism for the unimpaired ihnibition of the protease Val82Ala mutant is also suggested, based on energy calculations and analyses.
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Phytoremediation and biodegradation of PCB in soil
Ryšlavá, E. ; Krejčík, Zdeněk ; Macek, Tomáš ; Macková, M.
This is a part of the research work focused on study of possible mutual relationships of microorganisms and plants in PCBs metabolism. Polluted soil was obtained from a site which has been contaminated by PCBs for a long time. This soil was used for isolation of bacteria whixh potentially participate in biolodegradation of PCBs. The same contaminated soil was used for experiments under real conditions. Within six months three plant species were cultivated-tobaco (Nicotiana tabacum), black nighshade (Solanum nigrum) and alfalfa (Medicago sativa) - and decrease of PCBs was measured.
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