Surface labeling of key residues during assembly of the transmembrane pore formed by staphylococcal α‐hemolysin

FEBS Letters

Braha, Orit¹  Krishnasastry, Musti¹  Walker, Barbara¹  Bayley, Hagan¹ 

[1] Worcester Foundation for Experimental Biology, 222 Maple Avenue, Shrewsbury, MA 01545, USA

关键词: Surface labeling;    Sulfhydryl modification;    Gel-shift electrophoresis;    Pore-forming protein;    Membrane protein;    Toxin;   

DOI  :  10.1016/0014-5793(94)01240-7

学科分类：生物化学/生物物理

来源: John Wiley & Sons Ltd.

【 摘 要 】

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.

【 授权许可】

Unknown