Features and mechanism of localized enzyme-assisted self-assembly of peptides from unilamellar vesicles

Localized enzyme-assisted self-assembly (LEASA) has emerged as a powerful tool to generate peptide nanofibers from and within bacteria or cancer cell lines. This approach has led to promising developments in medical imaging, antimicrobial treatments and cancer therapies. Despite these achievements, the features of self-assembly processes localized near the plasma cell membranes and induced by enzymes are not easy to study since biological media compositions are complex and vary over time. From model systems based on giant and small unilamellar vesicles (GUV and SUV respectively) displaying phosphatases at the outer edge of their phospholipid’s membrane, we study the self-assembly of peptides triggered by an enzymatic dephosphorylation. Peptide nanofibers, generated in the close vicinity of GUVs, adsorb all around the membrane where enzymes are located. This process induces the formation of a transient permeability through the membrane without destroying the vesicles, as observed by confocal laser scanning microscopy (CLSM). A cryo-transmission electron microscopy (cryo-TEM) monitoring over time highlights a lag-time before the formation of nano-aggregates located all around SUVs, followed by a rapid formation of short nanofibers near or directly from vesicles. Thanks to the presence of enzymes located on the surface of the vesicles, the micrometer-long nanofibers both adhere and exert such a mechanical force on the spherical shape of vesicles that they deform them. Finally, based on classical molecular dynamics simulations, we propose a mechanism that accounts for all our experimental observations, rationalizing the LEASA process induced on the surface of phospholipid bilayers containing phosphatases.