FIGURE 6.

The anchor peptide is not sufficient for binding to TbpA. Preparations of His-tagged anchor peptide (amino acids 2–40) fused to intact TbpB, TbpB N-lobe, or Mbp were captured on an Ni-NTA resin. Holo (iron-loaded) pTf was added to one set of samples (+pTf) prior to the addition of untagged TbpA. After incubation overnight, the resins were collected by centrifugation and washed twice, and the final pellet was resuspended in SDS-PAGE buffer to elute bound proteins. The samples were applied to an SDS-PAGE gel and stained with Coomassie Blue protein stain. MWS, molecular weight standard.