Subunit stoichiometry of staphylococcal α-hemolysin in crystals and on membranes

A heptameric transmembrane pore

Eric Gouaux, Orit Braha, Michael R. Hobaugh, Langzhou Song, Stephen Cheley, Christopher Shustak, Hagan Bayley

Research output: Contribution to journalArticle

210 Citations (Scopus)

Abstract

Elucidation of the accurate subunit stoichiometry of oligomeric membrane proteins is fraught with complexities. The interpretations of chemical cross- linking, analytical ultracentrifugation, gel filtration, and low-resolution electron microscopy studies are often ambiguous. Staphylococcal α-hemolysin (αHL), a homooligomeric toxin that forms channels in cell membranes, was believed to possess six subunits arranged around a sixfold axis of symmetry. Here, we report that analysis of x-ray diffraction data and chemical modification experiments indicate that the αHL oligomer is a heptamer. Self- rotation functions calculated using x-ray diffraction data from single crystals of αHL oligomers show a sevenfold axis of rotational symmetry. The αHL pore formed on rabbit erythrocyte membranes was determined to be a heptamer by electrophoretic separation of αHL heteromers formed from subunits with the charge of wild-type αHL and subunits with additional negative charge generated by targeted chemical modification of a single- cysteine mutant. These data establish the heptameric oligomerization state of the αHL transmembrane pore both in three-dimensional crystals and on a biological membrane.

Original languageEnglish (US)
Pages (from-to)12828-12831
Number of pages4
JournalProceedings of the National Academy of Sciences of the United States of America
Volume91
Issue number26
DOIs
StatePublished - Dec 20 1994
Externally publishedYes

Fingerprint

Hemolysin Proteins
Membranes
X-Rays
Ultracentrifugation
Erythrocyte Membrane
Gel Chromatography
Cysteine
Electron Microscopy
Membrane Proteins
Cell Membrane
Rabbits

ASJC Scopus subject areas

  • Genetics
  • General

Cite this

Subunit stoichiometry of staphylococcal α-hemolysin in crystals and on membranes : A heptameric transmembrane pore. / Gouaux, Eric; Braha, Orit; Hobaugh, Michael R.; Song, Langzhou; Cheley, Stephen; Shustak, Christopher; Bayley, Hagan.

In: Proceedings of the National Academy of Sciences of the United States of America, Vol. 91, No. 26, 20.12.1994, p. 12828-12831.

Research output: Contribution to journalArticle

Gouaux, Eric ; Braha, Orit ; Hobaugh, Michael R. ; Song, Langzhou ; Cheley, Stephen ; Shustak, Christopher ; Bayley, Hagan. / Subunit stoichiometry of staphylococcal α-hemolysin in crystals and on membranes : A heptameric transmembrane pore. In: Proceedings of the National Academy of Sciences of the United States of America. 1994 ; Vol. 91, No. 26. pp. 12828-12831.
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