Limits...
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.

Show MeSH

Related in: MedlinePlus

Graphic analyses of chemicalcross-link data for different models.(a) Graphic analysis of the goodness of fit of the five models plusthe entire 30 ns trajectory to cross-link violation distances, numbervs sum, assuming a cross-linking distance cutoff of 23 Å. (b)Overlay of 41 chemical cross-linking data points onto a static contactmap plot of Cα distances of ≤23 Å for the 15 nsmodel. Small purple circles represent the positions of all potentialcross-links, red circles the most probable one-third of cross-links,yellow circles the median probable one-third of cross-links, and bluecircles the least probable one-third of cross-links. Red arrowheadsdenote four loops and turns numbered from the N-terminus and bluearrowheads the N- and C-termini. Two cross-links (182–195 and195–206) at turn/loop 4 are denoted with red bull’s-eyesand four cross-links of H5 to H10 (118–226, 118–239,133–239, and 140–239) with magenta bull’s-eyes,and the cross-link supporting an H5 hairpin (118–140) is denotedwith a blue bull’s-eye.
© Copyright Policy
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4263436&req=5

fig4: Graphic analyses of chemicalcross-link data for different models.(a) Graphic analysis of the goodness of fit of the five models plusthe entire 30 ns trajectory to cross-link violation distances, numbervs sum, assuming a cross-linking distance cutoff of 23 Å. (b)Overlay of 41 chemical cross-linking data points onto a static contactmap plot of Cα distances of ≤23 Å for the 15 nsmodel. Small purple circles represent the positions of all potentialcross-links, red circles the most probable one-third of cross-links,yellow circles the median probable one-third of cross-links, and bluecircles the least probable one-third of cross-links. Red arrowheadsdenote four loops and turns numbered from the N-terminus and bluearrowheads the N- and C-termini. Two cross-links (182–195 and195–206) at turn/loop 4 are denoted with red bull’s-eyesand four cross-links of H5 to H10 (118–226, 118–239,133–239, and 140–239) with magenta bull’s-eyes,and the cross-link supporting an H5 hairpin (118–140) is denotedwith a blue bull’s-eye.

Mentions: We applied twomethods to establish the goodness of fit of themodels with the cross-linking data. First, we used the sum of violationdistances algorithm recently described by Kalisman et al.42 to analyze the goodness of fit of differentmodels to cross-link violation distances. We assumed a maximal Cαdistance of linkable lysine residues of 23 Å, defined as 20 Å+ 3.0 Å, the estimated coordinate error for two mobile surfaceresidues.33 For each of the six models,the number of violations and the sum of minimal violations were plottedon the x- and y-axes, respectively(Figure 4a). In this plot, the initial crystalmodel is farthest from the origin, while the Pollard, Silva, and 10–20ns models, in rank order, fall progressively closer to the origin.The best fit for any model (i.e., the one falling closest to the origin)was that of the 15 ns model. For comparison, a full trajectory analysisof the 0–30 ns MD simulation at 500 K, termed the full trajectorymodel, is also shown in Figure 4a. It shouldbe noted that, in the latter analysis, every structure in the simulationwas considered and the minimum of each individual violation distancewas found among possibly different structures.


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)

Graphic analyses of chemicalcross-link data for different models.(a) Graphic analysis of the goodness of fit of the five models plusthe entire 30 ns trajectory to cross-link violation distances, numbervs sum, assuming a cross-linking distance cutoff of 23 Å. (b)Overlay of 41 chemical cross-linking data points onto a static contactmap plot of Cα distances of ≤23 Å for the 15 nsmodel. Small purple circles represent the positions of all potentialcross-links, red circles the most probable one-third of cross-links,yellow circles the median probable one-third of cross-links, and bluecircles the least probable one-third of cross-links. Red arrowheadsdenote four loops and turns numbered from the N-terminus and bluearrowheads the N- and C-termini. Two cross-links (182–195 and195–206) at turn/loop 4 are denoted with red bull’s-eyesand four cross-links of H5 to H10 (118–226, 118–239,133–239, and 140–239) with magenta bull’s-eyes,and the cross-link supporting an H5 hairpin (118–140) is denotedwith a blue bull’s-eye.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4263436&req=5

fig4: Graphic analyses of chemicalcross-link data for different models.(a) Graphic analysis of the goodness of fit of the five models plusthe entire 30 ns trajectory to cross-link violation distances, numbervs sum, assuming a cross-linking distance cutoff of 23 Å. (b)Overlay of 41 chemical cross-linking data points onto a static contactmap plot of Cα distances of ≤23 Å for the 15 nsmodel. Small purple circles represent the positions of all potentialcross-links, red circles the most probable one-third of cross-links,yellow circles the median probable one-third of cross-links, and bluecircles the least probable one-third of cross-links. Red arrowheadsdenote four loops and turns numbered from the N-terminus and bluearrowheads the N- and C-termini. Two cross-links (182–195 and195–206) at turn/loop 4 are denoted with red bull’s-eyesand four cross-links of H5 to H10 (118–226, 118–239,133–239, and 140–239) with magenta bull’s-eyes,and the cross-link supporting an H5 hairpin (118–140) is denotedwith a blue bull’s-eye.
Mentions: We applied twomethods to establish the goodness of fit of themodels with the cross-linking data. First, we used the sum of violationdistances algorithm recently described by Kalisman et al.42 to analyze the goodness of fit of differentmodels to cross-link violation distances. We assumed a maximal Cαdistance of linkable lysine residues of 23 Å, defined as 20 Å+ 3.0 Å, the estimated coordinate error for two mobile surfaceresidues.33 For each of the six models,the number of violations and the sum of minimal violations were plottedon the x- and y-axes, respectively(Figure 4a). In this plot, the initial crystalmodel is farthest from the origin, while the Pollard, Silva, and 10–20ns models, in rank order, fall progressively closer to the origin.The best fit for any model (i.e., the one falling closest to the origin)was that of the 15 ns model. For comparison, a full trajectory analysisof the 0–30 ns MD simulation at 500 K, termed the full trajectorymodel, is also shown in Figure 4a. It shouldbe noted that, in the latter analysis, every structure in the simulationwas considered and the minimum of each individual violation distancewas found among possibly different structures.

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