RESUMO
In both prokaryotes and eukaryotes, the survival at temperatures considerably exceeding the optimum is supported by intense synthesis of the so-called heat shock proteins (HSPs), which act to overcome the adverse effects of heat stress. Among mycoplasmas (class Mollicutes), which have significantly reduced genomes, only some members of the Acholeplasmataceae family possess small HSPs of the α-crystallin type. Overproduction of a recombinant HSP IbpA (Hsp20) from the free-living mycoplasma Acholeplasma laidlawii was shown to increase the resistance of Escherichia coli to short-term heat shock. It has been long assumed that IbpA prevents protein aggregation and precipitation thereby increasing viability of E. coli cells. Several potential target proteins interacting with IbpA under heat stress were identified, including biosynthetic enzymes, enzymes of energy metabolism, and components of the protein synthesis machinery. Statistical analysis of physicochemical properties indicated that IbpA interaction partners significantly differ in molecular weight, charge, and isoelectric point from other members of the E. coli proteome. Upon shortterm exposure to increased temperature, IbpA was found to preferentially interact with high-molecular weight proteins having a pI of about 5.1, significantly lower than the typical values of E. coli proteins.
Assuntos
Acholeplasma laidlawii/química , Proteínas de Bactérias/química , Escherichia coli/fisiologia , Proteínas de Choque Térmico Pequenas/química , Temperatura Alta , Proteínas Recombinantes/química , Estresse FisiológicoRESUMO
Cell cultures of higher organisms, especially cultures of human cells, are increasingly used in medical, pharmaceutical and scientific research. The main problem of cell cultures non-lethal hidden contamination by mycoplasmas, viruses and outsider cell lines. As an available and reliable method for monitoring the purity of the cell cultures, we offer to use PCR kits designed and officially used in clinical diagnostics. We have tested 50 human cell lines using commercial diagnostic systems for detection of papilloma viruses, herpes viruses, adenoviruses, Mycoplasma hominis and total bacterial mass. Contamination in tested cell lines was not found. In the case of cell lines that contain integrated parts of viral genomes, the presence of the respective DNA sequences was confirmed. The proposed diagnostic systems can be effectively used to control the purity of cell lines, for qualitative detection of possible contamination, as well as for quantitative evaluations with calculation of viral load like it is practiced in clinical diagnostics.
Assuntos
Infecções por Vírus de DNA , Vírus de DNA/genética , Infecções por Mycoplasma , Mycoplasma hominis/genética , Reação em Cadeia da Polimerase/métodos , Técnicas de Cultura de Células , Linhagem Celular Tumoral , Infecções por Vírus de DNA/diagnóstico , Infecções por Vírus de DNA/genética , Humanos , Infecções por Mycoplasma/diagnóstico , Infecções por Mycoplasma/genéticaRESUMO
FtsZ--a bacterial tubulin homolog--is one of the key bacterial division proteins, forming a contractile Z-ring at the midcell of dividing bacteria. In this work immunofluorescent labeling was used in conjunction with single-molecule localization microscopy (SMLM) to visualize native structures formed by FtsZ protein in Escherichia coli cells. This approach allowed the reorganization of FtsZ structures during cytokinesis to be visualized step-by-step. New data was obtained that support the hypothesis that the Z-ring is a spiral structure that constricts during division, assisting the formation of the septum between daughter cells.