INTERACTIONS OF DEDICATED EXPORT MEMBRANE-PROTEINS OF THE COLICIN-V SECRETION SYSTEM - CVAA, A MEMBER OF THE MEMBRANE-FUSION PROTEIN FAMILY, INTERACTS WITH CVAB AND TOLC

The antibacterial peptide toxin colicin V uses a dedicated signal sequ ence-independent system for its secretion in Escherichia coli and requ ires the products of three genes, cvaA, cvaB, and tolC, As a member of the membrane fusion protein family, CvaA is supposed to form a bridge that connects the inner and outer membranes via interaction with CvaB and TolC, respectively, In this study, we investigated the possible i nteraction of these proteins, When CvaA or CvaB was absent, the corres ponding amount of CvaB or CvaA, respectively, was decreased, and the a mounts of both proteins were reduced when TolC was depleted, Translati onal lacZ fusions showed that TolC did not affect the synthesis of eit her CvaA-beta-galactosidase or CvaB-beta-galactosidase, and CvaA or Cv aB did not affect the synthesis of CvaB-beta-galactosidase or CvaA-bet a-galactosidase, respectively, However, the stabilities of CvaA and Cv aB proteins were affected by the absence of one another and by that of TolC, The instability of CvaA was more severe in TolC-depleted cells than in CvaB-depleted cells, On the other hand, CvaB was less stable i n the absence of CvaA than in the absence of TolC, In addition, using a cross-linking reagent, we showed that CvaA directly interacts with b oth CvaB and TolC proteins, Taken together, these data support the hyp othesized structural role of CvaA in connecting CvaB and TolC.