DISTINCT REGIONS OF THE COLICIN-A TRANSLOCATION DOMAIN ARE INVOLVED IN THE INTERACTION WITH TOLA AND TOLB PROTEINS UPON IMPORT INTO ESCHERICHIA-COLI

Group A colicins need proteins of the Escherichia coli envelope Tol co mplex (TolA, TolB, TolQ and TolR) to reach their cellular target. The N-terminal domain of colicins is involved in the import process. The N -terminal domains of colicins A and E1 have been shown to interact wit h TolA, and the N-terminal domain of colicin E3 has been shown to inte ract with TolB. We found that a pentapeptide conserved in the N-termin al domain of all group A colicins, the 'TolA box', was important for c olicin A import but was not involved in the colicin A-TolA interaction . It was, however, involved in the colicin A-TolB interaction. The int eractions of colicin A N-terminal domain deletion mutants with TolA an d TolB were investigated. Random mutagenesis was performed on a constr uct allowing the colicin A N-terminal domain to be exported in the bac teria periplasm. This enabled us to select mutant protein domains unab le to compete with the wildtype domain of the entire colicin A for imp ort into the cells. Our results demonstrate that different regions of the colicin A N-terminal domain interact with TolA and TolB. The colic in A N-terminal domain was also shown to form a trimeric complex with TolA and TolB.