MEMBRANE TOPOLOGIES OF THE TOLQ AND TOLR PROTEINS OF ESCHERICHIA-COLI- INACTIVATION OF TOLQ BY A MISSENSE MUTATION IN THE PROPOSED 1ST TRANSMEMBRANE SEGMENT

The TolQ and TolR proteins of Escherichia coli are required for the up take of group A colicins and for infection by filamentous phages. Thei r topology in the cytoplasmic membrane was determined by cleavage with aminopeptidase K, proteinase K, and trypsin in spheroplasts and cell lysates. From the results obtained, it is proposed that the N terminus of TolQ is located in the periplasm and that it contains three transm embrane segments (residues 9 to 36, 127 to 159, and 162 to 191), a sma ll periplasmic loop, and two large portions in the cytoplasm. The N te rminus of TolR is located in the cytoplasm and is followed by a transm embrane segment (residues 21 to 40), and the remainder of the protein is located in the periplasm. A tolQ mutant, which rendered cells resis tant to group A colicins and sensitive to cholate, had alanine 13 repl aced by glycine and was lacking serine 14 in the first transmembrane s egment. The membrane topologies of TolQ and TolR are similar to those proposed for ExbB and ExbD, respectively which is consistent with the partial functional substitution between ExbB and TolQ and between ExbD and TolR. The amino acid sequences of these proteins display the high est homology in the transmembrane segments, which indicates that the m embrane-spanning regions play an important role in the activities of t he proteins.