Investigating the Interaction of Saposin C with POPS and POPC Phospholipids: A Solid-State NMR Spectroscopic Study

Abstract

The interaction of Saposin C (Sap C) with negatively charged phospholipids such as phosphatidylserine (PS) is essential for its biological function. In this study, Sap C (initially protonated in a weak acid) was inserted into multilamellar vesicles (MLVs) consisting of either 1-palmitoyl-2-oleoyl- sn-glycero-3-[phospho-L-serine] (negatively charged, POPS) or 1-palmitoyl-2-oleoyl- sn-glycero-3-phosphocholine (neutrally charged, POPC). The MLVs were then investigated using solid-state NMR spectroscopy under neutral pH (7.0) conditions. The 2H and 31P solid-state NMR spectroscopic data of Sap C-POPS and Sap C-POPC MLVs (prepared under the same conditions) were compared using the 2H order parameter profiles of the POPC-d 31 or POPS-d 31 acyl chains as well as the 31P chemical shift anisotropy width and 31P T 1 relaxation times of the phospholipids headgroups. All those solid-state NMR spectroscopic approaches indicate that protonated Sap C disturbs the POPS bilayers and not the POPC lipid bilayers. These observations suggest for the first time that protonated Sap C inserts into PS bilayers and forms a stable complex with the lipids even after resuspension under neutral buffer conditions. Additionally, 31P solid-state NMR spectroscopic studies of mechanically oriented phospholipids on glass plates were conducted and perturbation effect of Sap C on both POPS and POPC bilayers was compared. Unlike POPC bilayers, the data indicates that protonated Sap C (initially protonated in a weak acid) was unable to produce well-oriented POPS bilayers on glass plates at neutral pH. Conversely, unprotonated Sap C (initially dissolved in a neutral buffer) did not interact significantly with POPS phospholipids allowing them to produce well-oriented bilayers at neutral pH.