Functional characterization of a melittin analog containing a non‐natural tryptophan analog. Issue 4 (July 2015)

• Record Type: Journal Article
• Authors: Ridgway, Zachary; Picciano, Angela L.; Gosavi, Pallavi M.; Moroz, Yurii S.; Angevine, Christopher E.; Chavis, Amy E.; Reiner, Joseph E.; Korendovych, Ivan V.; Caputo, Gregory A.
• Is Part Of: Biopolymers. Volume 104:Issue 4(2015)
• Page Start: 384
• Page End: 394
• Publication Date: 2015-07
• Subjects: fluorescence -- tryptophan analogs -- antimicrobial peptides -- azulene; Biopolymers -- Periodicals; Peptides -- Periodicals; Spectrum analysis -- Periodicals; 572.33
• DOI: 10.1002/bip.22624
• Languages: English
• ISSNs: 0006-3525

Abstract

ABSTRACT: Tryptophan (Trp) is a naturally occurring amino acid, which exhibits fluorescence emission properties that are dependent on the polarity of the local environment around the Trp side chain. However, this sensitivity also complicates interpretation of fluorescence emission data. A non‐natural analogue of tryptophan, β‐(1‐azulenyl)‐L‐alanine, exhibits fluorescence insensitive to local solvent polarity and does not impact the structure or characteristics of several peptides examined. In this study, we investigated the effect of replacing Trp with β‐(1‐azulenyl)‐L‐alanine in the well‐known bee‐venom peptide melittin. This peptide provides a model framework for investigating the impact of replacing Trp with β‐(1‐azulenyl)‐L‐alanine in a functional peptide system that undergoes significant shifts in Trp fluorescence emission upon binding to lipid bilayers. Microbiological methods including assessment of the antimicrobial activity by minimal inhibitory concentration (MIC) assays and bacterial membrane permeability assays indicated little difference between the Trp and the β‐(1‐azulenyl)‐L‐alanine‐substituted versions of melittin. Circular dichroism spectroscopy showed both that peptides adopted the expected α‐helical structures when bound to phospholipid bilayers and electrophysiological analysis indicated that both created membrane disruptions leading to significant conductance increases across model membranes. Both peptides exhibited a marked protection of the respective fluorophores when bound to bilayers indicating a similar membrane‐bound topology. As expected, while fluorescence quenching and CD indicate the peptides are stably bound to lipid vesicles, the peptide containing β‐(1‐azulenyl)‐L‐alanine exhibited no fluorescence emission shift upon binding while the natural Trp exhibited >10 nm shift in emission spectrum barycenter. Taken together, the β‐(1‐azulenyl)‐L‐alanine can serve as a solvent insensitive alternative to Trp that does not have significant impacts on structure or function of membrane interacting peptides. © 2015 Wiley Periodicals, Inc. Biopolymers (Pept Sci) 104: 384–394, 2015.