FIG. 6.
The TolQ periplasmic loop L164 residue is required for TolQ-KT interaction. (A) Colicin spot assays using serial dilutions of colicin K (from top to bottom, 1,000, 100, 10, and 1 ng of colicins have been spotted) on tolQ cells producing HA epitope-tagged TolQ (WT) or cysteine variants (V159C to A168C). (B) Detergent-solubilized extracts of tolQ cells coproducing TolQHA or its Cys derivatives and the FLAG epitope-tagged colicin K N-terminal domain were subjected to immunoprecipitation using the anti-FLAG antibody. The total solubilized material (input [Inp]) and the precipitated material (IP) were loaded on a 12.5% acrylamide SDS-PAGE gel and immunodetected with the anti-HA (TolQHA) and anti-FLAG (KT) monoclonal antibodies. Immunodetected proteins are indicated on the right. Molecular mass markers (in kDa) are indicated on the left.