The proposed research will focus on helix-helix interactions, the connecting loops and RNA recognition. The protein to be used for the proposed studies is called Rop. The function of Rop is to regulate the copy number of the plasmid ColE1 in E. coli. Rop is attractive for a variety of reasons: it can be readily over expressed and purified in large amounts from E. coli; it has no co- factors, disulfide bonds or proline residues; its x-ray crystal structure is known at high resolution; its proton NMR spectrum has been assigned; its stability can be determined in vitro by heat or denaturant induced unfolding (monitored by circular dichroism or differential scanning calorimetry); in vitro gel shift assays, with single stem-loop pairs, can be used to determine RNA binding affinity; its in vivo function can be exploited. %%% As a result of the proposed research Dr. Regan will obtain some very interesting novel proteins and it will be important to obtain more detailed structural information about them.
