Reduced B subunit of heat-labile enterotoxin associates with membranes in vivo

The B subunit of heat-labile enterotoxin, a periplasmic protein of Escherichia coli has an internal disulfide bond that forms after the protein has been exported. The presence of 2.5 mM dithiothreitol. in the medium prevents the formation of the disulfide bond and this causes the protein to rapidly bind to membranes, preferentially but not exclusively to the cytoplasmic membrane. The binding is irreversible in vivo but chaotropic agents disrupt the association between the non-native B subunit and the membranes in vitro. The fact that the reduced B subunit binds to both the cytoplasmic and outer membranes that enclose the periplasm suggests that it is exported normally to the periplasm and then, because it is unable to form its native structure, adsorbs to membranes in the vicinity. This is confirmed by the finding that when synthesised by spheroplasts, in which the outer membrane is disrupted, the majority of reduced B subunit, which is not now confined in the periplasm, is exported to the medium and is not associated with membranes.