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The P-113 Fragment of Histatin 5 Requires a Specific Peptide Sequence for Intracellular Translocation in Candida albicans, Which Is Independent of Cell Wall Binding

Department of Oral Biology,¹ Restorative Dentistry, School of Dental Medicine, State University of New York at Buffalo, Buffalo, New York 14214²

Received 11 September 2007/ Returned for modification 22 October 2007/ Accepted 1 November 2007

The activity of histatin 5 (Hst 5) against Candida albicans is initiated through cell wall binding, followed by translocation and intracellular targeting. The C. albicans cell wall protein Ssa2 is involved in the transport of Hst 5 into cells as part of cell killing. P-113 (a 12-amino-acid candidacidal active fragment of Hst 5) and P-113Q2.10 (which is inactivated by a glutamine substitution of the Lys residues at positions 2 and 10) were compared for their levels of cell wall binding and intracellular translocation in Candida wild-type (wt) and ssa2 strains. Both P-113 and P-113Q2.10 bound to the walls of C. albicans wt and ssa2 cells, although the quantity of P-113Q2.10 in cell wall extracts was higher than that of P-113 in both strains. Increasing the extracellular NaCl concentration to 100 mM completely inhibited the cell wall association of both peptides, suggesting that these interactions are primarily ionic. The accumulation of P-113 in the cytosol of wt cells reached maximal levels within 15 min (0.26 µg/10⁷ cells), while ssa2 mutant cells had maximal cytosolic levels of less than 0.2 µg/10⁷ cells even after 30 min of incubation. Furthermore, P-113 but not P-113Q2.10 showed specific binding with a peptide array of C. albicans Ssa2p. P-113Q2.10 was not transported into the cytosol of either C. albicans wt or ssa2 cells, despite the high levels of cell wall binding, showing that the two cationic lysine residues at positions 2 and 10 in the P-113 peptide are important for transport into the cytosol and that binding and transport are independent functional events.

Published ahead of print on 12 November 2007.