The highly conserved protease, Lon, regulates a diverse set of physiological responses in Escherichia coli, including cell division, capsule production, plasmid stability and phage replication (corresponding Lon substrates: SulA, RcsA, CcdA, and lambda N, respectively). Lon protease has been identified in every organism examined thus far, including Gram positive and Gram negative bacteria, yeast, plants and homo sapiens. To date, very little is known about the mechanism of substrate recognition by Lon. In an attempt to better understand the Lon-substrate interaction, point mutations in lon were generated which affected its substrate specificity. One such mutant class (referred to as lon *) behaved phenotypically and biochemically as if it were a lon - mutant with respect to RcsA (mucoid), yet acted lon + with respect to SulA (resistant to DNA damaging agents). DNA sequence analysis identified a single base change, leading to a single amino acid change within a highly conserved domain of Lon protease. The objective of this research is to test the hypothesis that this highly conserved domain is involved in the recognition and capture of Lon's substrates. Specific amino acid residues in this highly conserved domain will be changed and the impact of this change on RcsA and SulA stability will be examined (monitored by mucoidyness and response to DNA damaging agents). Understanding the protein-protein interactions which contribute to substrate recognition and capture by a protease will provide the foundation with which to construct a plausible model describing how crucial regulatory proteins are selected for degradation. The knowledge gained from these studies will extend understanding as to how selective degradation impacts the cell's goal to survive and proliferate.
This POWRE award will facilitate the PI's reinvigorating her research at a critical juncture in her career, thus it fits the POWRE criteria.
