Abstract
Apolipoprotein A-I (apoA-I) is the major protein in high density lipoprotein (HDL). During lipid metabolism, apoA-I moves among HDL and triacylglycerol-rich lipoproteins. The main structure and the major lipid binding motif of apoA-I is the amphipathic α-helix. To understand how apoA-I behaves at hydrophobic lipoprotein interfaces, the interfacial properties of apoA-I and an amphipathic α-helical consensus sequence peptide (CSP) were studied at the triolein/water (TO/W) interface. CSP ((PLAEELRARLRAQLEELRERLG)<sub>2</sub>-NH<sub>2</sub>) contains two 22-residue tandem repeat sequences that form amphipathic α-helices modeling the central part of apoA-I. ApoA-I or CSP added into the aqueous phase surrounding a triolein drop lowered the interfacial tension (γ) of TO/W in a concentration- and time-dependent fashion. The γ<sub>TO/W</sub> was lowered ∼16 millinewtons (mN)/m by apoA-I at 1.4 × 10<sup>–6</sup> m and ∼15 mN/m by CSP at 2.6 × 10<sup>–6</sup> m. At equilibrium γ, both apoA-I and CSP desorbed from the interface when compressed and readsorbed when expanded. The maximum surface pressure CSP could withstand without being ejected (Π<sub>MAX</sub>) was 16 mN/m. The Π<sub>MAX</sub> of apoA-I was only 14.8 mN/m, but re-adsorption kinetics suggested that only part of the apoA-I desorbed at Π between 14.8 and 19 mN/m. However, above ∼19 mN/m (Π<sub>OFF</sub>) the entire apoA-I molecule desorbed into the water. ApoA-I was more flexible at the TO/W interface than CSP and showed more elasticity at oscillation periods 4–128 s even at high compression, whereas CSP was elastic only at faster periods (4 and 8 s) and moderate compression. Flexibility and surface pressure-mediated desorption and re-adsorption of apoA-I probably provides lipoprotein stability during metabolic-remodeling reactions in plasma.
Highlights
ApoA-I exists in at least three states during metabolism: lipid-poor or free in solution, bound to a discoidal nascent high density lipoprotein (HDL) adsorbed when expanded
At 7 ϫ 10Ϫ9 M, it started at about 5000 s and resulted in a retardation of the fall in ␥ with time. We suggest this might be related to a rearrangement in the triolein surface in the presence of consensus sequence peptide (CSP), because we did not see this discontinuity in the ␥-time curves of CSP adsorbed onto DD/W or A/W interfaces [28], nor in the Apolipoprotein A-I (apoA-I) ␥-time curves
ApoA-I is the major apolipoprotein of HDL and a critical factor in the formation of nascent phospholipid/cholesterol HDL through interaction with ABCA1 and perhaps ABCG-1 (ATP-binding cassette (ABC) transporter G-1) and G-4 [35] transporters
Summary
The CSP peptide ((PLAEELRARLRAQLEELRERLG)2-NH2), which contains two 22-amino acid tandem repeats derived from the consensus sequence of the tandem repeats of human apoA-I, apoA-IV, and apoE [28, 29], was synthesized at Quality Controlled Biochemical Inc., using a Biosearch9050 Plus Continuous Flow Synthesizer, and purified Ͼ97ϩ%. To measure the interfacial tension of the TO/W interface with CSP, varied amounts of peptide stocks were added to the aqueous phase to obtain different peptide concentrations (from 7 ϫ 10Ϫ9 M to 5 ϫ 10Ϫ6 M). For interfacial tension measurements on the TO/W interface, varied amounts of apoA-I stocks were added to the aqueous phase to obtain different protein concentrations (from 1.7 ϫ 10Ϫ8 M to 1.4 ϫ 10Ϫ6 M). The pH of the aqueous phase was kept at pH 7.4 with phosphate buffer (2 mM). The interfacial tension of the TO/W interface in the presence of different amounts of CSP or apoA-I in the aqueous phase was measured with an I. 8-l triolein drops were formed in gently stirred pH 7.4 phosphate buffer (6.0 ml) containing a given amount of apoA-I or CSP peptide. All experiments were carried out at 25 Ϯ 0.1 °C in a thermostated system
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