HELP! Extraction of periplasmic proteins.

schupp at nauvax.ucc.nau.edu schupp at nauvax.ucc.nau.edu
Tue Dec 20 09:35:50 EST 1994


Alex,
        You might try polymyxin B sulfate, a strongly cationic antibiotic.  It
has been shown to disrupt the outer membrane of gram- bacteria.  I use it to
perforate cells prior to a luciferase assay at a concentration of 12.5ug/ml final.  The disruption is almost instantaneous at room temp.  I can't say whether this will selectively release periplasmic proteins while retaining intracellular
ones.  You might try a range of concentrations and compare the extraction p
protein profiles.  Papers by Donohue-Rolfe (Infect. Immun.  39:270-274, 1983)
and Griffin (Infect. Immun.  40:425-428, 1983) suggest that periplasmic
proteins can be selectively released by polymyxin B and osmotic shock.

Jim Schupp



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