Structure and lipid interactions of an anti-inflammatory and anti-atherogenic 10-residue class G⁎ apolipoprotein J peptide using solution NMR

Abstract

The surprising observation that a 10 residue class G* peptide from apolipoprotein J, [113-122]apoJ, possesses anti-inflammatory and anti-atherogenic properties prompted us to delineate its structural characteristics in the presence of normal and oxidized lipid. Towards this, we have determined high resolution structure of [113-122]apoJ in solution using nuclear magnetic resonance (NMR) spectroscopy and studied its interaction with lipids, including oxidized lipids, using a number of biophysical methods. Circular dichroism and NMR studies established that in the presence of dodecylphosphocholine (DPC) micelle this peptide adopts amphipathic α helical structure. The observed Nuclear Overhauser effects indicate that the amphipathic helical structure of the peptide is stabilized by the N-terminal acetyl and C-terminal amide blocking groups. We used isothermal titration calorimetry to measure binding enthalpy of the peptide with DPC micelle, an oxidized lipid, 1-(palmitoyl)-2-(5-keto-6-octene-dioyl) phosphatidylcholine (KOdiA-PC), and the mixture of these two lipids (5mol% KOdiA-PC in DPC micelle). We find that the peptide binding with DPC micelle is associated with an enthalpy change (-16.75±0.16 Kcal/mol) much larger than that resulting from the binding with KodiA-PC (-3.67±0.13 Kcal/mol). Incorporation of a small amount of KOdiA-PC (5mol %) in DPC micelle also results in the lowering of peptide binding enthalpy (-13.43±0.18 Kcal/mol). These results are consistent with overall negative charge and altered conformational properties of oxidized sn-2 chain of KOdiA-PC. Our results have unambiguously established the amphipathic α helical structure of [113-122]apoJ peptide in the presence of DPC micelle as well as its ability to bind oxidized lipid. These in vitro results help explain the previously observed anti-inflammatory and anti-atherosclerotic properties of this peptide.

Sidedness of interfacial arginine residues and anti-atherogenicity of apolipoprotein A-I mimetic peptides

“…Peptides 4F, [K 4,15 >R]4F, and [K 9,13 >R]4F ( Fig. 1 ) were synthesized by the solid phase method as described earlier ( 11 ).…”

“…The overall lipid affi nity of an amphipathic helix is partly dependent upon its depth of lipid penetration ( 11,25 ). For many of the residues on the nonpolar face, depth of lipid penetration decreases in the following order: 4F>[K 4,15 >R]4F>[K 9,13 >R]4F ( Fig. 1B ).…”

“…The CD spectra were measured from 260 nm to 190 nm every 0.5 nm with 4 s averaging per point and a 2 nm bandwidth. The secondary structure of the peptide was analyzed as described earlier ( 13,14 ).…”

“…Peptide:POPC by others (6)(7)(8)(9)(10). Peptides [K 4,15 >R]4F and [K 9,13 >R]4F, with Arg residues substituted for Lys residues on the right and left side respectively ( Fig. 1A ), were compared for their ability to inhibit atherosclerosis in older female apoE null mice with established aortic atherosclerotic lesions.…”

“…1B ). For example, depth of lipid penetration of Val10 is 10.6Å, 10.0Å, and 9.0Å for 4F, [K 4,15 >R]4F, and [K 9,13 >R]4F, respectively ( Fig. 1B ).…”