title: A Systematic Investigation of Structure/Function Requirements for the Apolipoprotein A-I/Lecithin Cholesterol Acyltransferase Interaction Loop of High-density Lipoprotein.
abstract: The interaction of lecithin-cholesterol acyltransferase (LCAT) with apolipoprotein A-I (apoA-I) plays a critical role in high-density lipoprotein (HDL) maturation. We previously identified a highly solvent-exposed apoA-I loop domain (Leu(159)-Leu(170)) in nascent HDL, the so-called "solar flare" (SF) region, and proposed that it serves as an LCAT docking site (Wu, Z., Wagner, M. A., Zheng, L., Parks, J. S., Shy, J. M., 3rd, Smith, J. D., Gogonea, V., and Hazen, S. L. (2007) Nat. Struct. Mol. Biol. 14, 861-868). The stability and role of the SF domain of apoA-I in supporting HDL binding and activation of LCAT are debated. Here we show by site-directed mutagenesis that multiple residues within the SF region (Pro(165), Tyr(166), Ser(167), and Asp(168)) of apoA-I are critical for both LCAT binding to HDL and LCAT catalytic efficiency. The critical role for possible hydrogen bond interaction at apoA-I Tyr(166) was further supported using reconstituted HDL generated from apoA-I mutants (Tyr(166) → Glu or Asn), which showed preservation in both LCAT binding affinity and catalytic efficiency. Moreover, the in vivo functional significance of NO2-Tyr(166)-apoA-I, a specific post-translational modification on apoA-I that is abundant within human atherosclerotic plaque, was further investigated by using the recombinant protein generated from E. coli containing a mutated orthogonal tRNA synthetase/tRNACUA pair enabling site-specific insertion of the unnatural amino acid into apoA-I. NO2-Tyr(166)-apoA-I, after subcutaneous injection into hLCAT(Tg/Tg), apoA-I(-/-) mice, showed impaired LCAT activation in vivo, with significant reduction in HDL cholesteryl ester formation. The present results thus identify multiple structural features within the solvent-exposed SF region of apoA-I of nascent HDL essential for optimal LCAT binding and catalytic efficiency.
authors.affiliation: From the Department of Cellular and Molecular Medicine, Lerner Research Institute, and.
authors.forename: Xiaodong
authors.initials: X
authors.lastname: Gu
authors.affiliation: From the Department of Cellular and Molecular Medicine, Lerner Research Institute, and.
authors.forename: Zhiping
authors.initials: Z
authors.lastname: Wu
authors.affiliation: From the Department of Cellular and Molecular Medicine, Lerner Research Institute, and.
authors.forename: Ying
authors.initials: Y
authors.lastname: Huang
authors.affiliation: From the Department of Cellular and Molecular Medicine, Lerner Research Institute, and.
authors.forename: Matthew A
authors.initials: MA
authors.lastname: Wagner
authors.affiliation: From the Department of Cellular and Molecular Medicine, Lerner Research Institute, and.
authors.forename: Camelia
authors.initials: C
authors.lastname: Baleanu-Gogonea
authors.affiliation: the Department of Biochemistry and Biophysics, Oregon State University, Corvallis, Oregon 97331, and.
authors.forename: Ryan A
authors.initials: RA
authors.lastname: Mehl
authors.affiliation: From the Department of Cellular and Molecular Medicine, Lerner Research Institute, and.
authors.forename: Jennifer A
authors.initials: JA
authors.lastname: Buffa
authors.affiliation: From the Department of Cellular and Molecular Medicine, Lerner Research Institute, and.
authors.forename: Anthony J
authors.initials: AJ
authors.lastname: DiDonato
authors.affiliation: From the Department of Cellular and Molecular Medicine, Lerner Research Institute, and.
authors.forename: Leah B
authors.initials: LB
authors.lastname: Hazen
authors.affiliation: From the Department of Cellular and Molecular Medicine, Lerner Research Institute, and.
authors.forename: Paul L
authors.initials: PL
authors.lastname: Fox
authors.affiliation: From the Department of Cellular and Molecular Medicine, Lerner Research Institute, and the Department of Chemistry, Cleveland State University, Cleveland, Ohio 44115.
authors.forename: Valentin
authors.initials: V
authors.lastname: Gogonea
authors.affiliation: the Sections on Molecular Medicine, Department of Internal Medicine, Wake Forest School of Medicine, Winston-Salem, North Carolina 27157.
authors.forename: John S
authors.initials: JS
authors.lastname: Parks
authors.affiliation: From the Department of Cellular and Molecular Medicine, Lerner Research Institute, and.
authors.forename: Joseph A
authors.initials: JA
authors.lastname: DiDonato
authors.affiliation: From the Department of Cellular and Molecular Medicine, Lerner Research Institute, and the Department of Cardiovascular Medicine, Cleveland Clinic, Cleveland, Ohio 44195, hazens@ccf.org.
authors.forename: Stanley L
authors.initials: SL
authors.lastname: Hazen
mesh_terms: D000818:Animals; D016632:Apolipoprotein A-I / Q000737:chemistry*; D006801:Humans; D007700:Kinetics; D008075:Lipoproteins, HDL / Q000737:chemistry*; D008810:Mice, Inbred C57BL; D007862:Phosphatidylcholine-Sterol O-Acyltransferase / Q000737:chemistry*; D011485:Protein Binding; D054730:Protein Interaction Domains and Motifs
chemical_list: D016632:Apolipoprotein A-I; D008075:Lipoproteins, HDL; D007862:Phosphatidylcholine-Sterol O-Acyltransferase
keywords: LCAT; apolipoprotein; atherosclerosis; cholesterol; high-density lipoprotein (HDL); post-translational modification (PTM)