PROTEIN COMPLEX WITHIN ESCHERICHIA-COLI INNER MEMBRANE - TOLA N-TERMINAL DOMAIN INTERACTS WITH TOLQ AND TOLR PROTEINS

The TolA, TolB, TolQ, and TolR proteins are involved in maintaining th e integrity of the Escherichia coil outer membrane and in the import o f group A colicins and filamentous phage DNA. TolA, TolQ, and TolR are localized in the inner membrane while TolB is periplasmic, although a small amount of membrane-associated TolB is always found. In vivo cro ss-linking experiments with formaldehyde were performed in order to de termine the proteins interacting with TolA In wild-type strains, two s pecific complexes of 65 and 71 kDa, com prising TolA, were identified. These complexes were absent in a tolQ strain, while only the 65-kDa c omplex was absent in a tolR strain. When the tol strains were transfor med with plasmids encoding TolR or TolQ, the specific complexes were r estored. Moreover, immunoprecipitation experiments with the antiserum directed against TolA indicated that TolQ and TolR were co-immunopreci pitated with TolA after cross-linking. These data demonstrate that Tol A interacts directly with TolR and TolQ. Two truncated TolA proteins d evoid of either the C-terminal or the central domains of the protein w ere subjected to in vivo cross-linking. Since these two TolA derivativ es still formed specific complexes with TolR and TolQ, we concluded th at the TolA N-terminal domain interacted with these proteins.