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Tuning Liposome Membrane Permeability by Competitive Peptide Dimerization and Partitioning-Folding Interactions Regulated by Proteolytic Activity.

Lim SK, Sandén C, Selegård R, Liedberg B, Aili D - Sci Rep (2016)

Bottom Line: We describe a de novo designed membrane active peptide that partition into lipid membranes only when specifically and covalently anchored to the membrane, resulting in pore-formation.The effect can be regulated by proteolytic digestion of the inhibitory peptide by the matrix metalloproteinase MMP-7, an enzyme upregulated in many malignant tumors.This system thus provides a precise and specific route for tuning the permeability of lipid membranes and a novel strategy for development of recognition based membrane active peptides and indirect enzymatically controlled release of liposomal cargo.

View Article: PubMed Central - PubMed

Affiliation: Centre for Biomimetic Sensor Science, School of Materials Science and Engineering, Nanyang Technological University, Research Techno Plaza, 6th storey XFrontiers block, 50 Nanyang Drive, 637553 Singapore.

ABSTRACT
Membrane active peptides are of large interest for development of drug delivery vehicles and therapeutics for treatment of multiple drug resistant infections. Lack of specificity can be detrimental and finding routes to tune specificity and activity of membrane active peptides is vital for improving their therapeutic efficacy and minimize harmful side effects. We describe a de novo designed membrane active peptide that partition into lipid membranes only when specifically and covalently anchored to the membrane, resulting in pore-formation. Dimerization with a complementary peptide efficiently inhibits formation of pores. The effect can be regulated by proteolytic digestion of the inhibitory peptide by the matrix metalloproteinase MMP-7, an enzyme upregulated in many malignant tumors. This system thus provides a precise and specific route for tuning the permeability of lipid membranes and a novel strategy for development of recognition based membrane active peptides and indirect enzymatically controlled release of liposomal cargo.

No MeSH data available.


Related in: MedlinePlus

CD spectra of JR2KC (10 μM in PBS pH 7.4) in the absence (orange, dashed) and presence of POPC liposomes with 0 mol% (orange) and 5 mol% (blue) MPB-PE.The scrambled polypeptide (10 μM in PBS pH 7.4) shows no ordered secondary structure in absence (purple, dashed) nor in presence of POPC with 5 mol% MPB-PE (purple). Inset: Ratio of [θ]222 and [θ]208 for JR2KC (10 μM in PBS pH 7.4) over time in the presence of POPC liposomes with 0 mol% (orange) and 5 mol% (blue) MPB-PE.
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f3: CD spectra of JR2KC (10 μM in PBS pH 7.4) in the absence (orange, dashed) and presence of POPC liposomes with 0 mol% (orange) and 5 mol% (blue) MPB-PE.The scrambled polypeptide (10 μM in PBS pH 7.4) shows no ordered secondary structure in absence (purple, dashed) nor in presence of POPC with 5 mol% MPB-PE (purple). Inset: Ratio of [θ]222 and [θ]208 for JR2KC (10 μM in PBS pH 7.4) over time in the presence of POPC liposomes with 0 mol% (orange) and 5 mol% (blue) MPB-PE.

Mentions: The influence of the membrane interactions on the peptide secondary structure was investigated using circular dichroism (CD) spectroscopy. JR2KC was random coil at neutral pH, both in the absence and presence of maleimide-free (0% MPB-PE) liposomes (Fig. 3A). In the presence of liposomes with 5 mol% MPB-PE, CD spectra of JR2KC showed a significant amount of helicity, with minima at 208 and 222 nm and a mean residue ellipticity at 222 nm ([Θ]222) of approximately −15000° cm2 dmol−1. The initial transition from random coil to α-helical was rapid, within minutes, but the helicity continued to increase for about 10 minutes after addition of the peptide (Fig. 3B). The folding kinetics was thus on par with the rate of CF release (Supplementary Fig. 3). The folding of the peptides suggest that membrane anchoring induces partitioning of JR2KC into the hydrophobic core of the membrane, which promote induction of secondary structure via partitioning-folding coupling, as observed in many AMPs. The membrane permeabilizing effect was abolished when scrambling the primary sequence of JR2KC, keeping only the position of the cysteine residue, to form a non-amphipathic peptide (Fig. 2 and Supplementary Fig. 1). In contrast to JR2KC this peptide did not fold in the presence of MPB-PE containing liposomes (Fig. 3), clearly demonstrating that the increase in membrane permeability caused by JR2KC is both sequence-specific and folding dependent in addition to requiring anchoring of the peptide to the membrane. Dynamic light scattering (DLS) experiments showed only minor changes in liposome size (hydrodynamic radius) for liposomes with and without MPB-PE in the presence of JR2KC (Supplementary Fig. 5). The binding of the peptides to the liposomes did consequently not trigger formation of micelles, nor liposome aggregation.


Tuning Liposome Membrane Permeability by Competitive Peptide Dimerization and Partitioning-Folding Interactions Regulated by Proteolytic Activity.

Lim SK, Sandén C, Selegård R, Liedberg B, Aili D - Sci Rep (2016)

CD spectra of JR2KC (10 μM in PBS pH 7.4) in the absence (orange, dashed) and presence of POPC liposomes with 0 mol% (orange) and 5 mol% (blue) MPB-PE.The scrambled polypeptide (10 μM in PBS pH 7.4) shows no ordered secondary structure in absence (purple, dashed) nor in presence of POPC with 5 mol% MPB-PE (purple). Inset: Ratio of [θ]222 and [θ]208 for JR2KC (10 μM in PBS pH 7.4) over time in the presence of POPC liposomes with 0 mol% (orange) and 5 mol% (blue) MPB-PE.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4759693&req=5

f3: CD spectra of JR2KC (10 μM in PBS pH 7.4) in the absence (orange, dashed) and presence of POPC liposomes with 0 mol% (orange) and 5 mol% (blue) MPB-PE.The scrambled polypeptide (10 μM in PBS pH 7.4) shows no ordered secondary structure in absence (purple, dashed) nor in presence of POPC with 5 mol% MPB-PE (purple). Inset: Ratio of [θ]222 and [θ]208 for JR2KC (10 μM in PBS pH 7.4) over time in the presence of POPC liposomes with 0 mol% (orange) and 5 mol% (blue) MPB-PE.
Mentions: The influence of the membrane interactions on the peptide secondary structure was investigated using circular dichroism (CD) spectroscopy. JR2KC was random coil at neutral pH, both in the absence and presence of maleimide-free (0% MPB-PE) liposomes (Fig. 3A). In the presence of liposomes with 5 mol% MPB-PE, CD spectra of JR2KC showed a significant amount of helicity, with minima at 208 and 222 nm and a mean residue ellipticity at 222 nm ([Θ]222) of approximately −15000° cm2 dmol−1. The initial transition from random coil to α-helical was rapid, within minutes, but the helicity continued to increase for about 10 minutes after addition of the peptide (Fig. 3B). The folding kinetics was thus on par with the rate of CF release (Supplementary Fig. 3). The folding of the peptides suggest that membrane anchoring induces partitioning of JR2KC into the hydrophobic core of the membrane, which promote induction of secondary structure via partitioning-folding coupling, as observed in many AMPs. The membrane permeabilizing effect was abolished when scrambling the primary sequence of JR2KC, keeping only the position of the cysteine residue, to form a non-amphipathic peptide (Fig. 2 and Supplementary Fig. 1). In contrast to JR2KC this peptide did not fold in the presence of MPB-PE containing liposomes (Fig. 3), clearly demonstrating that the increase in membrane permeability caused by JR2KC is both sequence-specific and folding dependent in addition to requiring anchoring of the peptide to the membrane. Dynamic light scattering (DLS) experiments showed only minor changes in liposome size (hydrodynamic radius) for liposomes with and without MPB-PE in the presence of JR2KC (Supplementary Fig. 5). The binding of the peptides to the liposomes did consequently not trigger formation of micelles, nor liposome aggregation.

Bottom Line: We describe a de novo designed membrane active peptide that partition into lipid membranes only when specifically and covalently anchored to the membrane, resulting in pore-formation.The effect can be regulated by proteolytic digestion of the inhibitory peptide by the matrix metalloproteinase MMP-7, an enzyme upregulated in many malignant tumors.This system thus provides a precise and specific route for tuning the permeability of lipid membranes and a novel strategy for development of recognition based membrane active peptides and indirect enzymatically controlled release of liposomal cargo.

View Article: PubMed Central - PubMed

Affiliation: Centre for Biomimetic Sensor Science, School of Materials Science and Engineering, Nanyang Technological University, Research Techno Plaza, 6th storey XFrontiers block, 50 Nanyang Drive, 637553 Singapore.

ABSTRACT
Membrane active peptides are of large interest for development of drug delivery vehicles and therapeutics for treatment of multiple drug resistant infections. Lack of specificity can be detrimental and finding routes to tune specificity and activity of membrane active peptides is vital for improving their therapeutic efficacy and minimize harmful side effects. We describe a de novo designed membrane active peptide that partition into lipid membranes only when specifically and covalently anchored to the membrane, resulting in pore-formation. Dimerization with a complementary peptide efficiently inhibits formation of pores. The effect can be regulated by proteolytic digestion of the inhibitory peptide by the matrix metalloproteinase MMP-7, an enzyme upregulated in many malignant tumors. This system thus provides a precise and specific route for tuning the permeability of lipid membranes and a novel strategy for development of recognition based membrane active peptides and indirect enzymatically controlled release of liposomal cargo.

No MeSH data available.


Related in: MedlinePlus