A Small Peptide Localizes to a Distinct Region of the Cell Membrane By Sensing Curvature

Many cellular functions rely on the correct subcellular localization of proteins. These proteins often localize by recognizing a pre-localized binding partner. How, then, do the very first proteins localize to its correct destination? The small peptide SpoVM, a critical component in endospore assembly in Bacillus subtilis, was shown to localize to the nascent forespore by sensing positive (convex) curvature within the cell membrane. We investigate how this nanometer-scale peptide can recognize curvature on the micrometer scale, and elucidate the critical features of the 26-amino acid peptide for sensing membrane curvature. We first reconstituted membrane curvature on the micron scale in vitro by creating solid-supported lipid bilayers (SSLBs) on silica beads, and show SpoVM preferentially binds to membranes with higher degrees of curvature. Next, we developed a flow cytometry assay to quantify binding of SpoVM to SSLBs and show that preferential binding can be attributed to a higher kinetic rate of binding, coupled with binding cooperativity. Finally, through the rational design of SpoVM mutants, we show that the curvature-sensing ability of SpoVM is dependent on a critical proline residue at position 9, which delineates the peptide into a flexible N-terminus and an amphipathic alpha-helical C-terminus. As more proteins are being reported to localize to curved membranes, these findings may reveal a common mechanism by which proteins achieve preferential binding to membranes with positive curvature.