Many antimicrobial peptides permeabilize the bacterial cytoplasmic membrane
. However, it is unclear how membrane permeabilization and antimicrobial ac
tivity are related for distinct peptides. This study investigated the relat
ionship between Staphylococcus aureus membrane permeabilization and cell de
ath due to the following antistaphylococcal peptides: thrombin-induced plat
elet microbicidal protein 1 (tPMP-1), gramicidin D, and protamine. Isogenic
S. aureus strains ISP479C and ISP479R (tPMP-1 susceptible and resistant, r
espectively), were loaded with the fluorochrome calcein and exposed to a ra
nge of concentrations of each peptide. Flow cytometry was then used to moni
tor membrane permeabilization by quantifying the release of preloaded calce
in. Killing was determined by quantitative culture at time points simultane
ous to measurement of membrane permeabilization. Membrane permeabilization
and killing caused by tPMP-1 occurred in a time- and concentration-dependen
t manner, reflecting the intrinsic tPMP-1 susceptibilities of ISP479C and I
SP479R. In comparison, gramicidin D killed both S. aureus strains to equiva
lent extents in a concentration-dependent manner between 0.5 to 50 mug/ml,
but cell permeabilization only occurred at the higher peptide concentration
s (25 and 50 mug/ml). Protamine permeabilized, but did not kill, either str
ain at concentrations up to 10 mg/ml. Regression analyses revealed differen
t relationships between membrane permeabilization and staphylocidal activit
y for the distinct antimicrobial peptides. Taken together, these findings d
emonstrate that permeabilization, per se, does not invariably result in sta
phylococcal death due to distinct antimicrobial peptides. Thus, although ea
ch of these peptides interacts with the S. aureus cytoplasmic membrane, div
ersity exists in their mechanisms of action with respect to the relationshi
p between membrane permeabilization and staphylocidal activity.