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

Shadi Abu-Baker ^*, Xiaoyang Qi  and Gary A. Lorigan ^*

^* Department of Chemistry and Biochemistry, Miami University, Oxford, Ohio; and Division and Program of Human Genetics, Cincinnati Children's Hospital Medical Center and University of Cincinnati College of Medicine, Cincinnati, Ohio

Correspondence: Address reprint requests to Gary A. Lorigan, Tel.: 513-529-3338; Fax: 513-529-5715; E-mail: garylorigan{at}muhio.edu.

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-d₃₁ or POPS-d₃₁ acyl chains as well as the ³¹P chemical shift anisotropy width and ³¹P T₁ 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.