Abstract
Mature proteins can act as potential sources of encrypted bioactive peptides that, once released from their parent proteins, might interact with diverse biomolecular targets. In recent work we introduced a systematic methodology to uncover encrypted intragenic antimicrobial peptides (IAPs) within large protein sequence libraries. Given that such peptides may interact with membranes in different ways, resulting in distinct observable outcomes, it is desirable to develop a predictive methodology to categorize membrane active peptides and establish a link to their physicochemical properties. Building upon previous work, we explored the interaction of a range of IAPs with model membranes probed by differential scanning calorimetry (DSC) and circular dichroism (CD) techniques. The biophysical data were submitted to multivariate statistical methods and resulting peptide clusters were correlated to peptide structure and to their antimicrobial activity. A re-evaluation of the physicochemical properties of the peptides was conducted based on peptide cluster memberships. Our data indicate that membranolytic peptides produce characteristic thermal transition (DSC) profiles in model vesicles and that this can be used to categorize novel molecules with unknown biological activity. Incremental expansion of the model presented here might result in a unified experimental framework for the prediction of novel classes of membrane active peptides.
Highlights
Genomes encode many proteins with borderline aqueous solubility that may partition into hydrophobic media or phospholipid membranes as soon as they are encountered1
These Intragenic Antimicrobial Peptides (IAPs), along with six known antimicrobial peptides (AMPs) (DS0112, Mag-2a13, Nattererin-114, Ascaphin-0815, PS-216, HSP-044), were incubated separately with Large Unilamellar Vesicules (LUVs) comprising either dimyristoylphosphatidylcholine (DMPC) alone or 2:1 dimyristoylphosphatidylcholine:dimyristoylphosphatidylglycerol (DMPC:DMPG) mixtures, and the resulting peptide-phospholipid LUVs were evaluated by differential scanning calorimetry (DSC) and circular dichroism (CD)
DSC data were submitted to a principal component analysis (PCA) followed by hierarchical clustering analysis (HCA), revealing three peptide clusters
Summary
Genomes encode many proteins with borderline aqueous solubility that may partition into hydrophobic media or phospholipid membranes as soon as they are encountered. Our methodology relies in two complementary steps: (1) the filtering of genomes/protein collections using a bioinformatic tool, “Kamal”, which searches protein databases for fragments using a predefined set of physicochemical properties; and (2) an experimental classification tool to identify sets of IAPs which induce comparable disturbances in model phospholipid membranes. The latter was conceived to gain further insight in peptide-membrane interactions and to serve as a feedback to the selection process by refining the physicochemical parameters associated with a particular biological activity. Future expansion of the currently presented methodology will assist the discovery of protein fragments with cell-penetrating and preamyloid activities and may provide a unified experimental framework for membrane active molecules
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