Mechanisms of Selective Antimicrobial Activity of Gaegurin 4

Abstract

Gaegurin 4 (GGN4), an antimicrobial peptide isolated from a Korean frog, is five times more potent against Gram-positive than Gram-negative bacteria, but has little hemolytic activity. To understand the mechanism of such cell selectivity, we examined GGN4-induced K ＋ efflux from target cells, and membrane conductances in planar lipid bilayers. The K ＋ efflux from Gram-positive M. luteus (2.5 μg/ml) was faster and larger than that from Gram-negative E. coli (75 μg/ml), while that from RBC was negligible even at higher concentration (100 μg/ml). GGN4 induced larger conductances in the planar bilayers which were formed with lipids extracted from Gram-positive B. subtilis than in those from E. coli (p＜ 0.01), however, the effects of GGN4 were not selective in the bilayers formed with lipids from E. coli and red blood cells. Addition of an acidic phospholipid, phosphatidylserine to planar bilayers increased the GGN4-induced membrane conductance (p＜ 0.05), but addition of phosphatidylcholine or cholesterol reduced it (p＜ 0.05). Transmission electron microscopy revealed that GGN4 induced pore-like damages in M . luteus and dis-layering damages on the outer wall of E. coli. Taken together, the present results indicate that the selectivity of GGN4 toward Gram-positive over Gram-negative bacteria is due to negative surface charges, and interaction of GGN4 with outer walls. The selectivity toward bacteria over RBC is due to the presence of phosphatidylcholine and cholesterol, and the trans-bilayer lipid asymmetry in RBC. The results suggest that design of selective antimicrobial peptides should be based on the composition and topology of membrane lipids in the target cells.

Trichoderma viride cellulase induces resistance to the antibiotic pore-forming peptide alamethicin associated with changes in the plasma membrane lipid composition of tobacco BY-2 cells

“…The CM-induced changes in phospholipids and their corresponding fatty acids (Figure 7 Figure 8 ), suggest that the physical properties of the plasma membrane were altered, possibly sufficient to affect alamethicin insertion and pore formation. This agrees with that the conductance through pores made by the antimicrobial cationic peptide gaegurin 4 was larger in planar bilayers made of PE, PC and PS (80:10:10) compared to membranes composed of only PE and PC (80:20) [ 63 ]. A role of sterols with respect to alamethicin channel activity was shown with artificial membranes, i.e ., the presence of cholesterol increased the duration of the alamethicin pore in its open state, indicating a more efficient use of created pores, while the critical concentration of alamethicin needed for pore formation increased [ 64 , 65 ].…”

“…Oligomerisation and pore formation by Vibrio cholerae cytolysin also depended on the presence of cholesterol [ 66 ]. With gaegurin 4 [ 63 ], inclusion of cholesterol in planar lipid membranes acted opposite to PS, i.e ., it prevented channel formation. This deviates from the association of increased alamethicin resistance to decreased sterol levels (relative to fatty acids) observed with tobacco cells (Figure 7 ).…”

pH Dependent Antimicrobial Peptides and Proteins, Their Mechanisms of Action and Potential as Therapeutic Agents

“…Similar PE mediated mechanisms have been proposed to mediate the resistance of microbes to other AMPs [104,105], supporting the growing view that receptors could play a variety of roles in the biological activities of these peptides [23,98,106,107,108]. Esculentin-2EM (E2EM, previously gaegurin 4) is an α-helical peptide isolated from the frog, Glandirana emeljanovi (formerly Rana rugosa ) [109,110], that is able to kill protozoa, fungi, Gram-positive bacteria and Gram-negative bacteria [110,111,112]. E2EM possesses a C-terminal cyclic region stabilized by a disulphide bond (Rana box) that is conserved across many ranid AMPs and helps stabilize pore formation by the peptide thereby promoting its antimicrobial action (Figure 1) [109,110,112,113,114,115].…”

“…Esculentin-2EM (E2EM, previously gaegurin 4) is an α-helical peptide isolated from the frog, Glandirana emeljanovi (formerly Rana rugosa ) [109,110], that is able to kill protozoa, fungi, Gram-positive bacteria and Gram-negative bacteria [110,111,112]. E2EM possesses a C-terminal cyclic region stabilized by a disulphide bond (Rana box) that is conserved across many ranid AMPs and helps stabilize pore formation by the peptide thereby promoting its antimicrobial action (Figure 1) [109,110,112,113,114,115]. Several models have been proposed to represent pore formation by E2EM and the best, supported by experimental evidence, appear to be the toroidal pore and barrel stave mechanisms (Figure 1).…”

Dipeptides Inhibit Melanin Synthesis in Mel-Ab Cells through Down-Regulation of Tyrosinase

“…Synthetic peptides show wide-ranging activities, some with significant effects on inflammation, immunity, antioxidant response, resistance to infection and melanogenesis [ 8 - 10 ]. While peptide synthesis is relatively simple, synthetic peptides may be expensive, depending on the number and type of amino acids required [ 11 ].…”