Dissecting the roles of ParA ATP binding and hydrolysis in P1 plasmid partition

grantor: University of TorontoThe partition system of the P1 bacteriophage ensures proper segregation of daughter plasmids at each division of the host 'Escherichia coli ' cell. It consists of ParA, ParB, and a 'cis'-acting site called 'parS'. ParA regulates expression of the ' par' operon and has an undefined, yet indispensable role in partition. Nucleotide binding and hydrolysis mediate ParA's regulatory and partition functions. I made 'parA' mutants to dissect the roles of ATP binding and hydrolysis. All of the mutants were defective for partition but could be divided into two subtypes with respect to repressor activity: class I strong repressors and class II partial repressors. Biochemical data showed that while all of the mutant proteins possessed reduced ATPase activity, the class I ParA's bound specifically to the 'par' promoter in a nucleotide-dependent manner whereas the class II ParA's did not. My data demonstrate that ATP hydrolysis is not required for ParA repressor activity but is essential for partition.M.Sc