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An experimentally robust model of monomeric apolipoprotein A-I created from a chimera of two X-ray structures and molecular dynamics simulations.

Segrest JP, Jones MK, Shao B, Heinecke JW - Biochemistry (2014)

Bottom Line: Consequently, we combined these crystal structures into an initial model that was subjected to molecular dynamics simulations.We tested the initial and simulated models and the two previously published models in three ways: against two published data sets (domains predicted to be helical by H/D exchange and six spin-coupled residues) and against our own experimentally determined cross-linking distance constraints.We note that the best fit simulation model, superior by all tests to previously published models, has dynamic features of a molten globule with interesting implications for the functions of apoA-I.

View Article: PubMed Central - PubMed

Affiliation: Department of Medicine, Atherosclerosis Research Unit, and Center for Computational and Structural Dynamics, University of Alabama at Birmingham , Birmingham, Alabama 35294, United States.

ABSTRACT
High-density lipoprotein (HDL) retards atherosclerosis by accepting cholesterol from the artery wall. However, the structure of the proposed acceptor, monomeric apolipoprotein A-I (apoA-I), the major protein of HDL, is poorly understood. Two published models for monomeric apoA-I used cross-linking distance constraints to derive best fit conformations. This approach has limitations. (i) Cross-linked peptides provide no information about secondary structure. (ii) A protein chain can be folded in multiple ways to create a best fit. (iii) Ad hoc folding of a secondary structure is unlikely to produce a stable orientation of hydrophobic and hydrophilic residues. To address these limitations, we used a different approach. We first noted that the dimeric apoA-I crystal structure, (Δ185-243)apoA-I, is topologically identical to a monomer in which helix 5 forms a helical hairpin, a monomer with a hydrophobic cleft running the length of the molecule. We then realized that a second crystal structure, (Δ1-43)apoA-I, contains a C-terminal structure that fits snuggly via aromatic and hydrophobic interactions into the hydrophobic cleft. Consequently, we combined these crystal structures into an initial model that was subjected to molecular dynamics simulations. We tested the initial and simulated models and the two previously published models in three ways: against two published data sets (domains predicted to be helical by H/D exchange and six spin-coupled residues) and against our own experimentally determined cross-linking distance constraints. We note that the best fit simulation model, superior by all tests to previously published models, has dynamic features of a molten globule with interesting implications for the functions of apoA-I.

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Stability analysis ofthe apoA-I monomer core CH1, CH4, and CH5domains. (a) Root-mean-square fluctuation (rmsf) of all residues ofthe initial crystal model of apoA-I over the entire course of the30 ns MD simulation at 500 K. Red boxes show H/D helical regions 1,4, and 5 (residues 7–44, 81–115, and 147–178,respectively) that represent the core helical domains of the monomermodel. Colored circles (L22 and L163, yellow; G26 and S167, gray;E92, red; K96, blue; Y100, Y104, Y108, and Y166, magenta; L159, green)denote key residues in panel b that stabilize the crossover contactsamong the three core helical domains. (b) Relaxed-eye stereo viewof the CH1, CH4, and CH5 domains (transparent red ribbons identifiedby pink numerals) of the 15 ns model. Hydrophobic residues of thecore are shown as green sticks. G26 and S167 (gray space-filling model)form a hydrogen-bonded pair at the crossover between CH1 and CH5.E92 (red space-filling model), K96 (blue space-filling model), andY166 (magenta space-filling model) form a cation−π complexbetween CH4 and CH5. Y100, F104, and W108 (magenta, dotted space-fillingmodel–stick combination) form an aromatic cup around L159 (greensticks) between CH4 and CH5. L22 on CH1 (yellow space-filling model)packs tightly against the G26–S167 hydrogen bond. L163 on CH5(yellow space-filling model) is tightly packed into the angle betweenS167 and Y166. Y115 on CH4 associates with three surrounding hydrophobicresidues to help hold the upper ends of CH4 and CH5 together. Similarhydrophobic interactions occur between CH4 and CH5 at the bottom.
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fig7: Stability analysis ofthe apoA-I monomer core CH1, CH4, and CH5domains. (a) Root-mean-square fluctuation (rmsf) of all residues ofthe initial crystal model of apoA-I over the entire course of the30 ns MD simulation at 500 K. Red boxes show H/D helical regions 1,4, and 5 (residues 7–44, 81–115, and 147–178,respectively) that represent the core helical domains of the monomermodel. Colored circles (L22 and L163, yellow; G26 and S167, gray;E92, red; K96, blue; Y100, Y104, Y108, and Y166, magenta; L159, green)denote key residues in panel b that stabilize the crossover contactsamong the three core helical domains. (b) Relaxed-eye stereo viewof the CH1, CH4, and CH5 domains (transparent red ribbons identifiedby pink numerals) of the 15 ns model. Hydrophobic residues of thecore are shown as green sticks. G26 and S167 (gray space-filling model)form a hydrogen-bonded pair at the crossover between CH1 and CH5.E92 (red space-filling model), K96 (blue space-filling model), andY166 (magenta space-filling model) form a cation−π complexbetween CH4 and CH5. Y100, F104, and W108 (magenta, dotted space-fillingmodel–stick combination) form an aromatic cup around L159 (greensticks) between CH4 and CH5. L22 on CH1 (yellow space-filling model)packs tightly against the G26–S167 hydrogen bond. L163 on CH5(yellow space-filling model) is tightly packed into the angle betweenS167 and Y166. Y115 on CH4 associates with three surrounding hydrophobicresidues to help hold the upper ends of CH4 and CH5 together. Similarhydrophobic interactions occur between CH4 and CH5 at the bottom.

Mentions: Althoughwe performed the simulation at 500 K, we found littlechange in overall helicity between 10 and 30 ns (58 to 55%). We surmisedfrom the H/D data40 that the structure’score conformation remains relatively intact [red helical domains 1(residues 7–31), 4 (residues 84–120), and 5 (residues146–174) in Figure 5a]. Evidence ofcore stability is provided by the rmsd data (Figure S2 of the Supporting Information) and by the more extensivestability analyses shown in Figure 7. An rmsfplot of the fluctuation of each residue during the full simulation(Figure 7a) shows that core helix (CH) domains1, 4, and 5 (along with smaller H/D helical domains 2 and 3) are morestable than most of the remainder of the model.


An experimentally robust model of monomeric apolipoprotein A-I created from a chimera of two X-ray structures and molecular dynamics simulations.

Segrest JP, Jones MK, Shao B, Heinecke JW - Biochemistry (2014)

Stability analysis ofthe apoA-I monomer core CH1, CH4, and CH5domains. (a) Root-mean-square fluctuation (rmsf) of all residues ofthe initial crystal model of apoA-I over the entire course of the30 ns MD simulation at 500 K. Red boxes show H/D helical regions 1,4, and 5 (residues 7–44, 81–115, and 147–178,respectively) that represent the core helical domains of the monomermodel. Colored circles (L22 and L163, yellow; G26 and S167, gray;E92, red; K96, blue; Y100, Y104, Y108, and Y166, magenta; L159, green)denote key residues in panel b that stabilize the crossover contactsamong the three core helical domains. (b) Relaxed-eye stereo viewof the CH1, CH4, and CH5 domains (transparent red ribbons identifiedby pink numerals) of the 15 ns model. Hydrophobic residues of thecore are shown as green sticks. G26 and S167 (gray space-filling model)form a hydrogen-bonded pair at the crossover between CH1 and CH5.E92 (red space-filling model), K96 (blue space-filling model), andY166 (magenta space-filling model) form a cation−π complexbetween CH4 and CH5. Y100, F104, and W108 (magenta, dotted space-fillingmodel–stick combination) form an aromatic cup around L159 (greensticks) between CH4 and CH5. L22 on CH1 (yellow space-filling model)packs tightly against the G26–S167 hydrogen bond. L163 on CH5(yellow space-filling model) is tightly packed into the angle betweenS167 and Y166. Y115 on CH4 associates with three surrounding hydrophobicresidues to help hold the upper ends of CH4 and CH5 together. Similarhydrophobic interactions occur between CH4 and CH5 at the bottom.
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fig7: Stability analysis ofthe apoA-I monomer core CH1, CH4, and CH5domains. (a) Root-mean-square fluctuation (rmsf) of all residues ofthe initial crystal model of apoA-I over the entire course of the30 ns MD simulation at 500 K. Red boxes show H/D helical regions 1,4, and 5 (residues 7–44, 81–115, and 147–178,respectively) that represent the core helical domains of the monomermodel. Colored circles (L22 and L163, yellow; G26 and S167, gray;E92, red; K96, blue; Y100, Y104, Y108, and Y166, magenta; L159, green)denote key residues in panel b that stabilize the crossover contactsamong the three core helical domains. (b) Relaxed-eye stereo viewof the CH1, CH4, and CH5 domains (transparent red ribbons identifiedby pink numerals) of the 15 ns model. Hydrophobic residues of thecore are shown as green sticks. G26 and S167 (gray space-filling model)form a hydrogen-bonded pair at the crossover between CH1 and CH5.E92 (red space-filling model), K96 (blue space-filling model), andY166 (magenta space-filling model) form a cation−π complexbetween CH4 and CH5. Y100, F104, and W108 (magenta, dotted space-fillingmodel–stick combination) form an aromatic cup around L159 (greensticks) between CH4 and CH5. L22 on CH1 (yellow space-filling model)packs tightly against the G26–S167 hydrogen bond. L163 on CH5(yellow space-filling model) is tightly packed into the angle betweenS167 and Y166. Y115 on CH4 associates with three surrounding hydrophobicresidues to help hold the upper ends of CH4 and CH5 together. Similarhydrophobic interactions occur between CH4 and CH5 at the bottom.
Mentions: Althoughwe performed the simulation at 500 K, we found littlechange in overall helicity between 10 and 30 ns (58 to 55%). We surmisedfrom the H/D data40 that the structure’score conformation remains relatively intact [red helical domains 1(residues 7–31), 4 (residues 84–120), and 5 (residues146–174) in Figure 5a]. Evidence ofcore stability is provided by the rmsd data (Figure S2 of the Supporting Information) and by the more extensivestability analyses shown in Figure 7. An rmsfplot of the fluctuation of each residue during the full simulation(Figure 7a) shows that core helix (CH) domains1, 4, and 5 (along with smaller H/D helical domains 2 and 3) are morestable than most of the remainder of the model.

Bottom Line: Consequently, we combined these crystal structures into an initial model that was subjected to molecular dynamics simulations.We tested the initial and simulated models and the two previously published models in three ways: against two published data sets (domains predicted to be helical by H/D exchange and six spin-coupled residues) and against our own experimentally determined cross-linking distance constraints.We note that the best fit simulation model, superior by all tests to previously published models, has dynamic features of a molten globule with interesting implications for the functions of apoA-I.

View Article: PubMed Central - PubMed

Affiliation: Department of Medicine, Atherosclerosis Research Unit, and Center for Computational and Structural Dynamics, University of Alabama at Birmingham , Birmingham, Alabama 35294, United States.

ABSTRACT
High-density lipoprotein (HDL) retards atherosclerosis by accepting cholesterol from the artery wall. However, the structure of the proposed acceptor, monomeric apolipoprotein A-I (apoA-I), the major protein of HDL, is poorly understood. Two published models for monomeric apoA-I used cross-linking distance constraints to derive best fit conformations. This approach has limitations. (i) Cross-linked peptides provide no information about secondary structure. (ii) A protein chain can be folded in multiple ways to create a best fit. (iii) Ad hoc folding of a secondary structure is unlikely to produce a stable orientation of hydrophobic and hydrophilic residues. To address these limitations, we used a different approach. We first noted that the dimeric apoA-I crystal structure, (Δ185-243)apoA-I, is topologically identical to a monomer in which helix 5 forms a helical hairpin, a monomer with a hydrophobic cleft running the length of the molecule. We then realized that a second crystal structure, (Δ1-43)apoA-I, contains a C-terminal structure that fits snuggly via aromatic and hydrophobic interactions into the hydrophobic cleft. Consequently, we combined these crystal structures into an initial model that was subjected to molecular dynamics simulations. We tested the initial and simulated models and the two previously published models in three ways: against two published data sets (domains predicted to be helical by H/D exchange and six spin-coupled residues) and against our own experimentally determined cross-linking distance constraints. We note that the best fit simulation model, superior by all tests to previously published models, has dynamic features of a molten globule with interesting implications for the functions of apoA-I.

Show MeSH
Related in: MedlinePlus