Solution Structure of a Novel Tryptophan-Rich Peptide with Bidirectional Antimicrobial Activity

Abstract

Trp-rich antimicrobial peptides play important roles in the host innate defense mechanisms of many plants, insects, and mammals. A new type of Trp-rich peptide, Ac-KWRRWVRWI-NH 2 , designated Pac-525, was found to possess improved activity against both gram-positive and -negative bacteria. We have determined that the solution structures of Pac-525 bound to membrane-mimetic sodium dodecyl sulfate (SDS) micelles. The SDS micelle-bound structure of Pac-525 adopts an ␣-helical segment at residues Trp2, Arg3, and Arg4. The positively charged residues are clustered together to form a hydrophilic patch. The three hydrophobic residues Trp2, Val6, and Ile9 form a hydrophobic core. The surface electrostatic potential map indicates the three tryptophan indole rings are packed against the peptide backbone and form an amphipathic structure. Moreover, the reverse sequence of Pac-525, Ac-IWRVWRRWK-NH 2 , designated Pac-525 rev , also demonstrates similar antimicrobial activity and structure in membrane-mimetic micelles and vesicles. A variety of biophysical and biochemical methods, including circular dichroism, fluorescence spectroscopy, and microcalorimetry, were used to show that Pac-525 interacted strongly with negatively charged phospholipid vesicles and induced efficient dye release from these vesicles, suggesting that the antimicrobial activity of Pac-525 may be due to interactions with bacterial membranes.

Cationic Antimicrobial Peptides

“…Examples are: the hexapeptide lactoferricin [58], indolicidin [59], TP [60], Pac-525 [61], the short bovine bactenecins analogs [54], and synthetic linear RW-rich hexapeptides indentified from combinatorial library studies [62]. On the level of the lipid membrane, the W residue has a high propensity to insert into the membrane and partition near the membrane-water interface [63], while positively charged R residues, with hydrogen bonding properties, provide the basis for peptide interaction with the anionic components of bacterial membranes.…”

The Design of Bacteria Strain Selective Antimicrobial Peptides Based on the Incorporation of Unnatural Amino Acids

“…Other concentrations of LUVs used are noted in the text and resulted from the dilution of the stock solution. Large unilamellar vesicles (LUVs) were prepared by extrusion using a Mini-Extruder (Avanti Polar Lipid Inc) (Wei, 2006) (Hunter et al, 2005). The solution was extruded through a 100 nm pore size polycarbonate membrane 21 times.…”

“…Contributions due SUVs were eliminated by subtracting the lipid spectra of the corresponding peptide-free solutions. All analysis of CD spectra was conducted after smoothing (with a means-movement function) and conversion to molar ellipticity using the JASCO Spectra Analysis program (Russell et al, 2010) (Wei, 2006) (Bringezu et al, 2007). CD spectra that exhibited HT values greater than 400 were not used due to excessive light scattering and / or absorption.…”

“…Data was analyzed with Origin® software (version 7.0). ITC data collection was obtained in duplicate in an effort to ensure reproducibility (Wieprecht, Apostolov, Beyermann, et al, 2000) (Russell et al, 2010) (Wei, 2006) .…”

A survey of the year 2007 literature on applications of isothermal titration calorimetry

“…(viii) Protein/peptide-lipid [323][324][325][326][327][328][329][330][331][332][333][334] (ix) Nucleic acid-small ligands (drugs) [14,25,44,[335][336][337][338][339][340][341][342][343][344][345][346][347][348][349][350][351][352][353] (x) Enzyme kinetics [48,49,354,355].…”