This Article Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Services Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Reprints and Permissions Copyright Information Books from ASM Press MicrobeWorld Google Scholar Articles by Strömstedt, A. A. Articles by Malmsten, M. PubMed PubMed Citation Articles by Strömstedt, A. A. Articles by Malmsten, M.

 Previous Article  |  Next Article 

Antimicrob. Agents Chemother. doi:10.1128/AAC.00477-08
Copyright (c) 2008, American Society for Microbiology and/or the Listed Authors/Institutions. All Rights Reserved.

Strategies for improving proteolytic resistance of antimicrobial peptides - Variants of EFK17, an internal segment of LL-37

Department of Pharmacy, Uppsala University, Uppsala, Sweden; Department of Dermatology and Venereology, Lund University, Lund, Sweden

^* To whom correspondence should be addressed. Email: adam.stromstedt{at}farmaci.uu.se.

	Abstract

Methods for increasing proteolytic stability were evaluated for EFK17 (EFKRIVQRIKDFLRNLV), a new peptide sequence with antimicrobial properties derived from LL-37. EFK17 was modified by four d-enantiomer or tryptophan (W) substitutions at known protease cleavage sites, as well as by terminal amidation and acetylation. The peptide variants were studied in terms of proteolytic resistance, antibacterial potency, and cytotoxicity, but also regarding their adsorption at model lipid membranes, liposomal leakage generation, and secondary structure behavior. The W substitutions resulted in a marked reduction of proteolytic degradation caused by human neutrophil elastase, Staphylococcus aureus aureolysin, and V8 protease, but not of degradation caused by Pseudomonas aeruginosa elastase. For the former two endoproteases also amidation and acetylation of the terminals reduced proteolytic degradation, but only when in combination with W substitutions. The d-enantiomer substitutions rendered the peptides indigestible by all four proteases, however those peptides displayed minor antimicrobial potency. The W- and end-modified peptides, on the other hand, showed an increased bactericidal potency compared to the native peptide sequence, coupled with a moderate cytotoxicity that was largely absent in serum. The bactericidal, cytotoxic, and liposome lytic properties correlated with each other, as well as with the amount of peptide adsorbed at the lipid membrane and the extent of helix formation associated with the adsorption. The lytic properties of the W-substituted peptides were less impaired by increased ionic strength, presumably by a combination of W-mediated stabilization of the largely amphiphilic helix conformation and a non-electrostatic W affinity for the bilayer interface. Overall, W substitutions constitute an interesting means to reduce the proteolytic susceptibility of EFK17, while also improving antimicrobial performance.