Antimicrobial Peptide GL13K Is Effective in Reducing Biofilms of Pseudomonas aeruginosa

Abstract

Human parotid secretory protein (PSP; BPIF2A) is predicted to be structurally similar to bactericidal/permeability-increasing protein and lipopolysaccharide (LPS)-binding protein. Based on the locations of known antimicrobial peptides in the latter two proteins, potential active peptides in the PSP sequence were identified. One such peptide, GL13NH 2 (PSP residues 141 to 153) was shown previously to interfere with LPS binding and agglutinate bacteria without bactericidal activity. By introducing three additional positively charged lysine residues, the peptide was converted to the novel bactericidal cationic peptide GL13K (MIC for Pseudomonas aeruginosa, 8 g/ml [5.6 M]). We investigated the antibiofilm activity of GL13K against static, monospecies biofilms of P. aeruginosa PAO1. Two-hour exposure of a 24-h biofilm to 64 g/ml (44.8 M) GL13K reduced biofilm bacteria by 10 2 , and 100 g/ml (70 M) GL13K reduced bacteria by 10 3 . Similar results could be achieved on 48-h-old biofilms. Lower concentrations of GL13K (32 g/ml [22.4 M]) were successful in reducing biofilm cell numbers in combination with tobramycin. This combination treatment also achieved total eradication of the biofilm in a majority (67.5%) of tested samples. An alanine scan of GL13K revealed the importance of the leucine residue in position six of the peptide sequence, where replacement led to a loss of antibiofilm activity, whereas the impact of replacing charged residues was less pronounced. Bacterial metalloproteases were found to partially inactivate GL13K but not a D amino acid version of the peptide.

Antimicrobial Peptides: Mechanisms of Action and Resistance

“…The polymer matrix may cause electrostatic repulsion of cationic peptides or trap the peptides to prevent them from reaching the embedded bacteria. As a result, only a minority of known AMPs, including GL13K (Hirt and Gorr 2013;Chen et al 2014), are also known to be active against bacterial biofilms (Di Luca et al 2015). Since biofilms are difficult to eradicate once they are established, recent strategies have focused on peptides that are specifically designed to prevent biofilm formation or eliminate existing biofilms (e.g., BAR and 1018; Daep et al 2010;de la Fuente-Núñez et al 2016).…”

“…Such AMPs include P113, which was derived from histatin 5 (Rothstein et al 2001);hLf1-11, from lactoferrin (Godoy-Gallardo et al 2014); and GL13K, from parotid secretory protein (BPIFA2; Abdolhosseini et al 2012;Balhara et al 2013;Hirt and Gorr 2013; Table 1). The goal for these peptides is to achieve strong antibacterial activity with low toxicity to mammalian cells and low ability to induce resistance in bacteria.…”

“…Substituting some or all amino acids in an AMP by D-amino acids has been used to increase resistance to proteolytic degradation of AMPs, including DJK-5, DJK-6, M33, and GL13K (Falciani et al 2012;Hirt and Gorr 2013;de la Fuente-Núñez et al 2015).…”

Antibacterial activity and cytocompatibility of an implant coating consisting of TiO₂ nanotubes combined with a GL13K antimicrobial peptide

“…It also adopts a favorable secondary structure in accordance with its surrounding environment to develop relatively hydrophilic or hydrophobic properties. 29 The minimal inhibitory concentration of GL13K is as low as 5-10 µg mL -1 against P. aeruginosa and E. coli, 30 and GL13K-modified Ti surfaces are antimicrobial against P. gingivalis. 16 Like other cationic AMPs, the cationic portions of GL13K interact with negatively charged bacterial membranes, and subsequently permeate or translocate across the membranes to disrupt their barrier function, 29,31 which forms the basis of their antimicrobial activities.…”

The dimerization of parotid secretory protein.

“…There is not a lot of research on the function of PSP, however, we do know that PSP binds to Phosphatidylinositol (3,4)Bisphosphate [37]. Recent research also shows that PSP binds to the sugars of lipopolysaccarides (LPS) [48] and can reduce biofilms confirming antimicrobial activity. With the understanding that PSP binds as a dimer we take a closer look at the different mutations of PSP and how it interacts with PIPs.…”