A Novel Linear Amphipathic β-Sheet Cationic Antimicrobial Peptide with Enhanced Selectivity for Bacterial Lipids

Jack Blazyk,Russell Wiegand,Jason Klein,Janet Hammer,Richard M Epand,Raquel F Epand,W Lee Maloy,U Prasad Kari

doi:10.1074/jbc.m102865200

Abstract

All known naturally occurring linear cationic peptides adopt an amphipathic alpha-helical conformation upon binding to lipids as an initial step in the induction of cell leakage. We designed an 18-residue peptide, (KIGAKI)3-NH2, that has no amphipathic character as an alpha-helix but can form a highly amphipathic beta-sheet. When bound to lipids, (KIGAKI)3-NH2 did indeed form a beta-sheet structure as evidenced by Fourier transform infrared and circular dichroism spectroscopy. The antimicrobial activity of this peptide was compared with that of (KIAGKIA)3-NH2, and it was better than that of GMASKAGAIAGKIAKVALKAL-NH2 (PGLa) and (KLAGLAK)3-NH2, all of which form amphipathic alpha-helices when bound to membranes. (KIGAKI)3-NH2 was much less effective at inducing leakage in lipid vesicles composed of mixtures of the acidic lipid, phosphatidylglycerol, and the neutral lipid, phosphatidylcholine, as compared with the other peptides. However, when phosphatidylethanolamine replaced phosphatidylcholine, the lytic potency of PGLa and the alpha-helical model peptides was reduced, whereas that of (KIGAKI)3-NH2 was improved. Fluorescence experiments using analogs containing a single tryptophan residue showed significant differences between (KIGAKI)3-NH2 and the alpha-helical peptides in their interactions with lipid vesicles. Because the data suggest enhanced selectivity between bacterial and mammalian lipids, linear amphipathic beta-sheet peptides such as (KIGAKI)3-NH2 warrant further investigation as potential antimicrobial agents.

Highlights

A diverse collection of host defense peptides discovered in a wide range of species shares the common characteristics of a net positive charge and the ability to form amphipathic structures
Our results show that KIGAKI does adopt a ␤-sheet conformation when bound to lipids and is comparable in antimicrobial activity to KIAGKIA and KLAGLAK
The amount of helical structure in KIAGKIA and KLAGLAK is approximately the same, whereas the helical content of KIGAKI is slightly lower under these conditions

Summary

TABLE I Amino acid sequences of antimicrobial peptides

GIGKFLHSAKKPGKAFVGEIMNS - NH2 GMASKAGAIAGKIAKVALKAL - NH2 KLASKAGKIAGKIAKVALKAL - NH2 KIAGKIAKIAGKIAKIAGKIA - NH2 KIAGKIAKWAGKIAKIAGKIA - NH2 KLAGLAKKLAGLAKKLAGLAK - NH2 KLAGLAKKWAGLAKKLAGLAK - NH2 KIGAKIKIGAKIKIGAKI - NH2 KIGAKIKWGAKIKIGAKI - NH2 signed a new peptide that can form a highly amphipathic ␤-sheet rather than an ␣-helix. This 18-residue peptide contains the hexameric repeat KIGAKI (Table I). The three model peptides, KIAGKIA, KLAGLAK, and KIGAKI, possess equal charge (ϩ7) and nearly equal mean hydrophobicity values. Our results show that KIGAKI does adopt a ␤-sheet conformation when bound to lipids and is comparable in antimicrobial activity to KIAGKIA and KLAGLAK. KIGAKI appears to possess greater selectivity for bacterial versus mammalian lipids as compared with the ␣-helical peptides tested

EXPERIMENTAL PROCEDURES

RESULTS

DISCUSSION

TABLE II Antimicrobial and hemolytic activities

KIAGKIA KLAGLAK KIGAKI