Structural changes in staphylococcal α-hemolysin (αHL) that occur during oligomerization and pore formation on membranes have been examined by using a simple gel-shift assay to determine the rate of modification of key single-cysteine mutants with the hydrophilic sulfhydryl reagent, 4-acetamido-4′-((iodoacetyl)amino)stilbene-2,2′-disulfonate (IASD). The central glycine-rich loop of αHL lines the lumen of the transmembrane channel. A residue in the loop remains accessible to IASD after assembly, in keeping with the ability of the pore to pass molecules of 1̃000 Da. By contrast, residues near the N-terminus, which are critical for pore function, become deeply buried during oligomerization, while a residue at the extreme C-terminus increases in reactivity after assembly, consistent with a location in the part of the pore that projects from the surface of the lipid bilayer. © 1994.
Krishnasastry, M., Walker, B., Braha, O., & Bayley, H. (1994). Surface labeling of key residues during assembly of the transmembrane pore formed by staphylococcal α-hemolysin. FEBS Letters, 356(1), 66–71. https://doi.org/10.1016/0014-5793(94)01240-7