Membrane insertion characteristics of the various transmembrane domains of the Escherichia coli TolQ protein.

The Escherichia coli TolQ protein is a 230-amino acid integral cytoplasmic membrane protein required for the import of group A colicins, for infection by the filamentous phage, and for maintenance of the integrity of the bacterial envelope. TolQ is a polytopic protein with three membrane-spanning regions. The first membrane-spanning region has a 19-residue periplasmic NH2-terminal tail, while the second and third membrane-spanning segments are separated by a short 17-amino acid periplasmic loop. To study the membrane assembly of TolQ, fusions of different membrane-spanning regions were examined for their ability to insert in the absence of functional SecA or the membrane potential. Fusions containing the first membrane-spanning region plus the adjacent cytoplasmic domain and a construct containing the "hairpin loop," formed by the second and third membrane-spanning regions, insert in the absence of functional SecA. The fusion containing the second and third membrane-spanning regions required the membrane potential for insertion while the first membrane-spanning region was able to insert even in the absence of a membrane potential. Taken together, these results suggest that insertion of intact TolQ is not dependent on the Sec system, but does require the membrane potential.