ISSN 1214-0287 (on-line), ISSN 1214-021X (printed)|
J Appl Biomed
Volume 8 (2010), No 3, p 151-158
DNA replication of plasmids in Gram-negative bacteria has been an object of study at CIB-CSIC for nearly 30 years. We have been focused on the enterobacterial antibiotic resistance factor R1 (1981-1992) and the pPS10 replicon from the phytopathogen Pseudomonas savastanoi (since 1984). Our group has used multidisciplinary (genetic, biochemical and biophysical-structural) approaches to unravel the molecular mechanism for the activation of RepA. Rep-type plasmidic proteins are either transcriptional repressors or replication initiators/inhibitors, depending on their association state (dimers vs. monomers) and targeting of alternative (operator or iteron) DNA sites. We discovered that allosteric DNA-binding remodels the structure of RepA N-terminal domain (WH1), transforming alpha-helical portions into beta-strands. This precisely tunes the distances between the DNA reading heads in WH1 and the C-terminal domain (WH2), to match the target operator or iteron sequences. We have recently moved into engineering such structural transformation in RepA-WH1 to build-up synthetic protein devices that allow for customized ligand (DNA)-promoted amyloidogenesis. Our basic studies on plasmid DNA replication are relevant for settling the bases of a minimalist bacterial model to tackle transmissible amyloid proteinopathies and are a valuable tool for bottom-up synthetic biology.
plasmid replication; protein amyloids; protein-DNA interactions; RepA protein; synthetic biology
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