Caracterização de cepas de Escherichia coli contendo diferentes alelos rpoS. / Characterization of Escherichia coli strains carrying different rpos alleles.
AUTOR(ES)
Heloisa Filus Galbiati
FONTE
IBICT - Instituto Brasileiro de Informação em Ciência e Tecnologia
DATA DE PUBLICAÇÃO
12/08/2011
RESUMO
Escherichia coli can be found in many different habitats and has to be prepared to survive and grow under unfavorable conditions. Bacteria adaptation to different growth conditions requires an efficient control of gene expression. One of the primary forms of gene expression control is the competition between different sigma factors for the binding to the core RNA polymerase. d70 is the most abundant sigma factor and participates in the transcription of most E. coli genes. dS is the second one in importance and recognizes promoters of genes related to the general stress response. The rpoS gene, which encodes dS, is highly polymorphic and acquires mutations very often. The transition C®T at position 97 in the rpoS ORF results in a stop codon TAG (amber). Due to the presence of an alternative Shine-Dalgarno and a translation initiation codon at position 157 a truncated RpoS protein that is partially functional is translated. One of the stress situations that dS is activated is the starvation for inorganic phosphate. Phosphate limitation also triggers the activation of the PHO regulon, whose genes are predominantly transcribed by d70. In the present study, the effect of the truncated version of RpoS on the expression of d70 dependent genes (lacZ, phoA e pstS) and dS dependent genes (osmY e proU) was tested. Bacteria were also assayed for sensitivity to oxidative, osmotic and cold stress. The profile of partial activity described for the truncated RpoS could be observed in some cases, while in others the behavior of this allele resembled the rpoS null mutant. In parallel, an E. coli strain which suppresses the amber mutation in RpoS, resulting in the expression of a normal protein was also tested. A band correponding to the truncated RpoS could not be detected in immunoblots probably due to inefficient translation from the alternative Shine-Dalgarno. To improve the detection of RpoS, a highly immunogenic SPA tag was inserted in the C-terminal region of the protein.
ASSUNTO(S)
escherichia coli escherichia coli ativação enzimática bacterial genetics enzyme activation genetic mutation genética bacteriana mutação genética rna polimerases rna polymerases
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