NMR structures and mutational analysis of the two peptides constituting the bacteriocin plantaricin S

Bie Ekblad,Per Eugen Kristiansen

doi:10.1038/s41598-019-38518-6

Abstract

The structure of the individual peptides of the two-peptide bacteriocin plantaricin S, an antimicrobial peptide produced by a Lactobacillus plantarum strain, has been determined in DPC micelles. The two peptides of plantaricin S, Pls-α and Pls-β, form an α-helix from and including residue 8 to 24 with a less structured region around residue 16-19 and an amphiphilic α-helix from and including residue 7 to 23, respectively. Activity assays on single amino acid-substituted GxxxG and GxxxG-like motifs show that substituting the Ser and Gly residues in the G9xxxG13 motif in Pls-α and the S17xxxG21 motif in Pls-β reduced or drastically reduced the antimicrobial activity. The two-peptide bacteriocin muricidin contains GxxxG-like motifs at similar positions and displays 40-50% amino acid identity with plantaricin S. Activity assays of combinations of the peptides that constitute the bacteriocins plantaricin S and muricidin show that some combinations are highly active. Furthermore, sequence alignments show that the motifs important for plantaricin S activity align with identical motifs in muricidin. Based on sequence comparison and activity assays, a membrane-inserted model of plantaricin S in which the two peptides are oriented antiparallel relative to each other and where the GxxxG and GxxxG-like motifs important for activity come close in space, is proposed.

Highlights

In view of the dramatic increase in antibiotic-resistant bacteria, the development of new antimicrobial drugs and food preservatives that kill and/or prevent growth of pathogenic bacteria is of increasing importance[1,2]
The antimicrobial activity of the two-peptide bacteriocins plantaricin JK and lactococcin G depends on a putative amino acid transporter[17,18] and a protein involved in cell wall synthesis[14], respectively
circular dichroism (CD) spectroscopy on plantaricin EF, plantaricin JK and lactococcin G have shown that when the peptides of two-peptide bacteriocins were mixed prior to the addition of liposomes they became more structured than the sum of the individual parts[23,24]

Summary

Introduction

In view of the dramatic increase in antibiotic-resistant bacteria, the development of new antimicrobial drugs and food preservatives that kill and/or prevent growth of pathogenic bacteria is of increasing importance[1,2]. Plantaricin S consists of the 27 residue Pls-α peptide and the 26 residue Pls-β peptide and is produced by Lab. plantarum strains involved in olive fermentation[9,10,11]. CD studies have revealed that the plantaricin S peptides are partly α-helical in 50% TFE/water and exhibited an additional α-helical structuring of 5% when combined[12] These results show that the peptides constituting a two-peptide bacteriocin can interact in vitro without the presence of a receptor[23,24]. All class IIb bacteriocins characterized so far contain one or several GxxxG or GxxxG-like motifs where x is any amino acid flanked by small residues such as Gly, Ser and Ala[8]. The peptides of muricidin have neither been characterized, synthesized nor assayed to determining antimicrobial activity

Methods

Results

Conclusion