Navegando por Palavras-chave "Csm"

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    Expressão, purificação, cristalização e análise preliminar de dados cristalográficos da proteína csm4 de thermotoga marítima
    (Universidade Federal de São Paulo (UNIFESP), 2015-11-30) Villegas, Nadia Valeria Gavilan [UNIFESP]; Alfonso, Martin Rodrigo Alejandro Wurtele [UNIFESP]; http://lattes.cnpq.br/0625673643875019; Universidade Federal de São Paulo (UNIFESP)
    Bacteria and Archaea have developed a defense system against exogenous DNA and RNA, called CRISPR (Clustered Regularly Intespaced Short Palindromic Repeats), which consists of an operon composed of regions of repetitions (repeats) identical to each other, separated by variable regions (spacers) from invading nucleic acids, along with associated proteins (Cas proteins, CRISPR associated proteins) forming an adaptive and heritable immune system. The main objective of this project was to clone, express and purify in recombinant form the Csm4 protein from T. maritima MSB8, and to characterize structurally this protein. This protein is part of a marginally studied complex of Cas ribonucleoproteins termed the Csm complex, which is involved in the targetting of exogenous DNA from bacteriophages and horizontal transfer of nucleic acids. The protein was recombinantly produced in E. coli and purified by affinity chromatography and gel filtration. After concentration, the protein was crystallized in space group P422 with unit cell a=67,5 b=67,5 c=84,8 Å. Data obtained showed that the native crystal diffracts to 2,5 Å at a wavelength of 1,458 Å. In order to solve the structure by MIR other crystals of derivatized protein with platinum and iodide diffracted up to 3,49 and 4,02 Å at wavelengths of 1,06007 and 1,54160 Å, respectively. Additionally, we incorporated selenomethionine into the protein to be able to solve the structure by the MAD (Multi-wavelength Anomalous Diffraction) methodology. These results can serve as a basis for implementing future projects of rational and experimental design of inhibitors in order to prevent the proliferation of these organisms.