An apoA-I mimetic peptide containing a proline residue has greater in vivo HDL binding and anti-inflammatory ability than the 4F peptide

Geoffrey D. Wool^*, Tomas Vaisar, Catherine A. Reardon^* and Godfrey S. Getz^*^,1

^* Department of Pathology, University of Chicago, Chicago, IL
Department of Medicine, University of Washington, Seattle, WA

¹ To whom correspondence should be addressed. e-mail: g-getz{at}uchicago.edu

Modifying apolipoprotein (apo) A-I mimetic peptides to includea proline-punctuated ${alpha}$ -helical repeat increases their anti-inflammatoryproperties as well as allows better mimicry of full-length apoA-Ifunction. This study compares the following mimetics, eitheracetylated or biotinylated (b): 4F (18mer) and 4F-proline-4F(37mer, Pro). b4F interacts with both mouse HDL (moHDL) andLDL in vitro. b4F in vivo plasma clearance kinetics are notaffected by mouse HDL level. Administration of biotinylatedpeptides to mice demonstrates that b4F does not associate withlipoproteins smaller than LDL in vivo, though it does associatewith fractions containing free hemoglobin (Hb). In contrast,bPro specifically interacts with HDL. b4F and bPro show oppositebinding responses to HDL by surface plasmon resonance. Administrationof acetylated Pro to apoE^–/– mice significantlydecreases plasma serum amyloid A levels, while acetylated 4Fdoes not have this ability. In contrast to previous reportsthat inferred that 4F associates with HDL in vivo, we systematicallyexamined this potential interaction and demonstrated that b4Fdoes not interact with HDL in vivo but rather elutes with Hb-containingplasma fractions. bPro, however, specifically binds to moHDLin vivo. In addition, the number of amphipathic ${alpha}$ -helices andtheir linker influences the anti-inflammatory effects of apoA-Imimetic peptides in vivo.

Supplementary key words apolipoprotein A-I • high density lipoprotein • synthetic peptides • cholesterol • inflammation • oxidation

Abbreviations: apo, apolipoprotein; AUC, area under the curve; b4F, biotinylated 4F; bPro, biotinylated Pro; DMF, dimethylformamide; ECL, enhanced chemiluminescence; FPLC, fast protein liquid chromatography; Hb, hemoglobin; HDL-C, HDL cholesterol; HRP, horseradish peroxidase; huHDL, human HDL; IP, intraperitoneal; MDA, malondialdehyde; moHDL, mouse HDL; moLDL, mouse LDL; Pro, 4F-proline-4F; RU, response unit; SAA, serum amyloid A; SPR, surface plasmon resonance; TBARS, thiobarbituric acid-reactive substance; TC, total cholesterol; TEP, 1,1,3,3-tetraethoxypropane

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