Regulatory mechanism of membrane protein production in an EPA-producing bacterium, Shewanella livingstonensis Ac10
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چکیده
A psychrotrophic Gram-negative bacterium, Shewanella livingstonensis Ac10, inducibly produces eicosapentaenoic acid (EPA) as a component of membrane phospholipids at low temperatures. We disrupted the genes required for EPA biosynthesis and found that the levels of the outer membrane porin homolog, Omp417, were markedly decreased in the EPA-less mutant (ΔEPA). To examine the effects of EPA on the folding of Omp417, in vitro refolding of recombinant Omp417 was carried out with liposomes in the presence or absence of EPA-containing phospholipids (EPA-PLs). Tryptophan (Trp) fluorescence dynamics of Omp417 reconstituted into liposomes with or without EPA-PLs demonstrated that the presence of EPA-PLs did not affect the local environments of Omp417 Trp residues. This result suggests that EPA-PLs are not involved in the folding of this protein at low temperatures. On the other hand, the transcription of omp417 was suppressed in the ΔEPA mutant, and the amount of omp417 transcript in the ΔEPA mutant was less than 2% of that in the wild type strain. To analyze the effects of EPA-PLs on omp417 expression, exogenous supplementation of EPA to ΔEPA cells and rescue of Δorf2 cells, a gene-disrupted mutant of a phosphopantetheinyl transferase required for the de novo synthesis of EPA, by using an orf2-expression vector were performed. Although these treatments restored EPA-PLs in the ΔEPA mutant, the transcriptional defect was not restored. These results suggest that the suppression of the transcription of omp417 is not due to the lack of EPA, but due to the insertion of a knockout plasmid for EPA-biosynthesis genes into the chromosome.
منابع مشابه
Purification and Characterization Strain Ac10: Gene Cloning and Enzyme Shewanella the Psychrotrophic Bacterium Cold-Active Serine Alkaline Protease from
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تاریخ انتشار 2016