Characterization of histatin 5 with respect to amphipathicity, hydrophobicity, and effects on cell and mitochondrial membrane integrity excludes a candidacidal mechanism of pore formation
Ej. Helmerhorst et al., Characterization of histatin 5 with respect to amphipathicity, hydrophobicity, and effects on cell and mitochondrial membrane integrity excludes a candidacidal mechanism of pore formation, J BIOL CHEM, 276(8), 2001, pp. 5643-5649
Histatin 5 is a 24-residue peptide from human saliva with antifungal proper
ties. We recently demonstrated that histatin 5 translocates across the yeas
t membrane and targets to the mitochondria, suggesting an unusual antifunga
l mechanism (Helmerhorst, E. J., Breeuwer, P., van 't Hof, W., Walgreen-Wet
erings, E., Oomen, L. C. J. M., Veerman, E. C. I., Nieuw Amerongen, k V., a
nd Abee, T. (1999) J. Biol. Chem. 274, 7286-7291). The present study used s
pecifically designed synthetic analogs of histatin 5 to elucidate the role
of peptide amphipathicity, hydrophobicity, and the propensity to adopt alph
a -helical structures in relation to membrane permeabilization and fungicid
al activity. Studies included circular dichroism measurements, evaluation o
f the effects on the cytoplasmic transmembrane potential and on the respira
tion of isolated mitochondria, and analysis of the peptide hydrophobicity/a
mphipathicity relationship (Eisenberg, D. (1984) Annu. Reu. Biochem. 53, 59
5-623). The 14-residue synthetic peptides used were dh-5, comprising the fu
nctional domain of histatin 5, and dhvar1 and dhvar4, both designed to maxi
mize amphipathic characteristics. The results obtained show that the amphip
athic analogs exhibited a high fungicidal activity, a high propensity to fo
rm an alpha -helix, dissipated the cytoplasmic transmembrane potential, and
uncoupled the respiration of isolated mitochondria, similar to the pore-fo
rming peptide PGLa ((P) under bar eptide with N-terminal Glycine and C-term
inal Leucine-amide). In contrast, histatin 5 and dh-5 showed fewer or none
of these features. The difference in these functional characteristics betwe
en histatin 5 and dh5 on the one hand and dhvar1, dhvar4, and PGLa on the o
ther hand correlated well with their predicted affinity for membranes based
on hydrophobicity/amphipathicity analysis. These data indicate that the sa
livary protein histatin 5 exerts its antifungal function through a mechanis
m other than pore formation.