Crystal structure of C-terminal truncated apolipoprotein A-I reveals the assembly of high density lipoprotein (HDL) by dimerization

Abstract

Apolipoprotein A-I (apoA-I) plays important structural and functional roles in plasma high density lipoprotein (HDL) that is responsible for reverse cholesterol transport. However, a molecular understanding of HDL assembly and function remains enigmatic. The 2.2-Å crystal structure of Δ(185-243)apoA-I reported here shows that it forms a half-circle dimer. The backbone of the dimer consists of two elongated antiparallel proline-kinked helices (five AB tandem repeats). The N-terminal domain of each molecule forms a four-helix bundle with the helical C-terminal region of the symmetry-related partner. The central region forms a flexible domain with two antiparallel helices connecting the bundles at each end. The two-domain dimer structure based on helical repeats suggests the role of apoA-I in the formation of discoidal HDL particles. Furthermore, the structure suggests the possible interaction with lecithin-cholesterol acyltransferase and may shed light on the molecular details of the effect of the Milano, Paris, and Fin mutations.

FIGURE 1.

Overall structure of Δ(185–243)apoA-I dimer. A, illustrations of the helix distribution from sequence analysis, consensus sequence model analysis, Δ(185–243)apoA-I crystal structure, and hydrogen exchange of plasma apoA-I and Δ(1–43)apoA-I crystal structure. Exon-3-encoded region (residues 1–43) is green. Exon-4 -encoded region (44–243) is ice-blue. Consensus sequence peptide (CSP) A and B homology sequences are in purple and cyan, respectively. Prolines are labeled in yellow. Five AB repeats are labeled with black dotted lines with arrows. B, overall structure of Δ(185–243)apoA-I monomer. C, two different views showing the homodimer with dimerization interface composed of two antiparallel five AB repeats. The N-terminal helix bundles are connected by the central segment hinge. Each region is colored to correspond to A. D, temperature factor distribution. The orange dotted circles show the most flexible regions.

FIGURE 2.

Structure of Δ(185–243)apoA-I dimer consists of N-terminal helix bundles and central segment hinge. N-terminal helix bundles of Δ(185–243)apoA-I dimer are stabilized by two aromatic clusters (N and C) and two π-cation interactions (N and C).

FIGURE 3.

Salt bridge interactions in monomer and between monomers. A, illustration of the potential salt bridges in the CSP-AB model and observed salt bridges in the five AB repeats from the monomer of the crystal structure on helix wheel diagrams. Salt bridges between i,i + 4 are connected with electric blue dotted lines with arrows. Salt bridges between i,i + 3 are connected with cyan dotted lines with arrows. Residues labeled with red star form salt bridge triads. Unusual salt bridges are connected with red dotted lines with arrows. Negatively charged residues are red; positively charged residues are blue; hydrophobic residues are white; prolines are yellow; neutral residues are light green, and histidine residues are blue and white. B, two major (N and C) salt bridge networks consisting of salt bridges between monomers and within the monomer hold the dimer of the crystal structure together. Arg¹⁷¹ and Arg¹⁵¹ labeled by red dotted lines are the positions corresponding to the Milano and Paris mutation, respectively. C, H6(AB4) and H4(AB2) region is the possible LCAT interaction region. Surface colored with residue charge is shown to identify possible charged residues that are responsible for the formation of salt bridges with LCAT.

FIGURE 4.

Surface properties of the Δ(185–243)apoA-I crystal structure. A, possible lipid entrance tunnel at the N-terminal helix bundle with surface colored according to the residue hydrophobicity. Enlarged figure shows the direct access to the C π-cation from the tunnel with the surface colored according to the residue charge. B, inside and outside view of the amphipathic tunnel formed by the central AB repeats (H5/H5) with surface colored according to the residue hydrophobicity or charge. The amphipathic central tunnel has a hydrophilic outside surface composed of charged residues and a hydrophobic inside surface of hydrophobic residues with Ala¹³⁰ in the center of the tunnel forming a possible portal for the transportation of lipid by LCAT.

FIGURE 5.

Stabilization of first 43 residues for apoA-I and possible monomer conformation in solution. A, stabilization of the first 43 residues for apoA-I in solution and a possible mechanism for unhinging of the N-terminal helix bundle upon lipid binding. B, possible Δ(185–243)apoA-I monomer. The two conformational states interconvert dependent on protein concentration with the central AB repeats (H5) folding or unfolding.

FIGURE 6.

Five AB repeats form backbone and dimerization interface in the Δ(185–243)apoA-I and Δ(1–43)apoA-I crystal structures. A, illustrations of the dimerization interface of Δ(185–243)apoA-I and Δ(1–43)apoA-I crystal structure. Exon-3-encoded region (residues 1–43) is green. Exon-4-encoded region (residues 44–243) is ice blue. Consensus sequence peptide (CSP) A and B homology sequences are in purple and cyan, respectively. Prolines are labeled in yellow. Red dotted region shows the five AB repeat dimerization interface. B, comparing the dimerization interfaces of Δ(185–243)apoA-I and Δ(1–43) apoA-I crystal structures.

FIGURE 7.

Discoidal HDL particles formation mechanism and size variation. A, possible mechanism of discoidal HDL particle formation from monomer apoA-I in solution through three states. B, HDL models based on the crystal structure resemble different sizes of discoidal HDL particles.