Research on Mechanism of Activating a Promoter by Bent DNA and it's Application

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Previously we showed that the expression of a Clostridium perfringens phospholipase C gene (plc) is activated by promoter upstream phased A-tracts in a low temperature-dependent manner. In this research we characterize the interaction between the α subunit of C. perfringens RNA polymerase and the phased A-tracts. Hydroxyl radical footprinting and fluorescence polarization assaying revealed that the α subunit binds to the minor grooves of the phased A- tracts through its C-terminal domain with increased affinity at low temperature. The result provides a molecular mechanism underlying the activation of the plc promoter by phased A-tractsIn order to clarify the contribution of the C-terminal domain of the α subunit on plc promoter activation, we tried to reconstitute of C. perfringens RNA polymerase, lacking the C-terminal domain. But we could not get any active RNA polymerase reconstituted with recombinant α, β, β', and o subunits of C. perfringens