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Why Is the C-terminus of Aβ(1-42) More Unfolded than That of Aβ(1-40)? Clues from Hydrophobic Interaction Liang Shen, Hong-Fang Ji,* and Hong-Yu Zhang* Shandong ProVincial Research Center for Bioinformatic Engineering and Technique, Center for AdVanced Study, Shandong UniVersity of Technology, Zibo 255049, People’s Republic of China. ReceiVed: August 11, 2007
Aβ(1-40) and Aβ(1-42) are the main forms of amyloid β (Aβ) peptides in the brain of Alzheimer’s patients; however, the latter possesses much stronger aggregation and deposition propensity than the former, which is partially attributed to the more unfolded C-terminus of Aβ(1-42) than that of Aβ(1-40). To explore the physical basis underlying the different dynamic behaviors of both Aβ peptides, parallel molecular dynamics (MD) simulations on Aβ(1-40) and Aβ(1-42) were performed to investigate their thermal unfolding processes. It is revealed that the addition of residues 41 and 42 in Aβ(1-42) disrupts the C-terminal hydrophobic core, which triggers the unraveling of the C-terminal helix of Aβ(1-42). This conclusion is supported by the MD simulation on the I41A mutant of Aβ(1-42), in which the C-terminal helix possesses relatively higher conformational stability than that of wild type Aβ(1-42) owing to the change in hydrophobic interaction patterns.
Introduction Alzheimer’s disease (AD), characterized by progressive memory loss, decline in language skills, and other cognitive impairments, has been the most common form of dementia among individuals over 65 years of age.1,2 Although the etiology of AD is not very clear, the amyloid β (Aβ) peptide fibril formation and deposition is widely considered as a central event in the pathogenetic process of AD.3,4 The Aβ(1-40) and Aβ(1-42) peptides, derived from proteolysis of amyloid precursor protein (APP), have been found to be the main forms of Aβ.5 Although Aβ(1-40) and Aβ(1-42) possess identical amino acid sequence (except that the latter has an additional two residues (IA) at the C-terminus), numerous in vitro and in vivo studies indicated that both peptides hold different aggregation and deposition properties.6-9 For example, Aβ(1-42) aggregates much faster than Aβ(1-40) in vitro6,7 and is more toxic than Aβ(1-40).8,9 A recent study employing transgenic mice revealed that Aβ(1-42) is sufficient for amyloid deposition in brain, while amyloid plaques are not formed in the case of high expression of Aβ(1-40).10 Since the polypeptide conformational changes from native state to unfolded state play critical roles in the amyloid formation,11-13 the different aggregation and deposition propensities of both peptides may be understood in terms of the more unfolded C-terminus of Aβ(1-42) than that of Aβ(1-40), as revealed by circular dichroism (CD) and nuclear magnetic resonance (NMR) experiments.14,15 However, the physical basis underlying the different dynamic behaviors of both Aβ peptides remains obscure, which stimulated our interest in employing molecular dynamics (MD) technique to explore why the C-terminus of Aβ(1-42) is more unfolded than that of Aβ(1-40). To avoid obtaining nonrepresentative results, the simulations for Aβ(1-40) and Aβ(1-42) were performed in parallel five times, respectively. The simulations were calculated at 400 K to accelerate the thermal unfolding processes * To whom correspondence should be addressed. Phone: (86) 5332780271. Fax: (86) 533-2780271. E-mail:
[email protected] (H.-F.J.);
[email protected] (H.-Y.Z.).
Figure 1. Evolution of RMSD of all atoms position during 5-ns simulation of Aβ(1-40) and Aβ(1-42) at 300 K (average of 5 parallel simulations) and 400 K (average of 5 parallel simulations), with respect to the initial structure. (a) Aβ(1-40) at 300 K; (b) Aβ(1-42) at 300 K; (c) I41A mutant of Aβ(1-42) at 300 K; (d) Aβ(1-40) at 400 K; (e) Aβ(1-42) at 400 K; (f) I41A mutant of Aβ(1-42) at 400 K.
of the two peptides. As a control, the native simulations at 300 K were also performed. Computational Methods The available NMR structure (protein data bank entry 1IYT)16 was used as starting structure of Aβ(1-42), while the initial structure of Aβ(1-40) was derived from excising the last two residues (IA) of Aβ(1-42). The I41A mutant of Aβ(1-42) was constructed through replacing Ile at position 41 by Ala using a “biopolymer” module of the InsightII program package.17 All the simulated structures were immersed in two layers (20 Å for the inner and 15 Å for the outer) of explicit water molecules. The inner layer was dynamic, while the outer layer was static and served as a solvent boundary to prevent the escape of the inner-layer water molecules. The models were minimized by 1000 conjugate gradient steps, heated from 2 to 300 K during
10.1021/jp7108237 CCC: $40.75 © 2008 American Chemical Society Published on Web 02/16/2008
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Figure 2. Snapshots of Aβ(1-40), Aβ(1-42), and the I41A mutant of Aβ(1-42) simulations extracted from the MD trajectories at different time scales at 400 K. Red, R-helix; green, random coil. As the five parallel simulations give very similar results, only one simulation snapshot is presented.
35 ps at temperature increment of 50 K per 5 ps, and kept at 300 K with 20 ps using the constant pressure and temperature algorithm. The velocity verlet integrator was used with an integration step of 2 fs. The production MD phase was carried out at 300 and 400 K, respectively, using 2-fs time steps and a 9.5-Å cutoff. The consistent valence force field (CVFF)18-21 was used in all of the calculations. Structures were saved every 0.5 ps for a total of 6000 snapshots for each trajectory. All of the NVT MD simulations were performed by a Discover module of InsightII program package,17 which has been widely employed in the MD simulation studies.22-25 According to the previous findings,26 a small number (5-10) of simulations are sufficient to capture the average properties of the unfolding pathways. Thus, to ensure the repeatability of the simulation results, a parallel 5-ns simulation for five Aβ(1-40), five Aβ(1-42), and five I41A mutants of Aβ(1-42) were performed. The simulation timescale is sufficient to reach the equilibrium states of the three peptides. All of the calculations were performed on a SGI Origin 350 server. Results and Discussion The root-mean-square deviations (RMSDs) of all atoms position relative to the initial energy-minimized conformation
of Aβ(1-40) and Aβ(1-42) at 300 and 400 K were calculated and shown in Figure 1. It can be seen that both Aβ(1-40) and Aβ(1-42) are rather stable during the native-state (300 K) simulations, while at 400 K, both Aβ(1-40) and Aβ(1-42) get unstable as indicated by the increased RMSDs. However, the RMSDs of Aβ(1-40) and Aβ(1-42) are close to each other, indicating that both peptides possess comparable overall stabilities, which is consistent with recent experimental findings.14,15 Moreover, the accelerated unraveling of both peptides from 300 to 400 K may account, at least in part, for the experimental observation that the Aβ aggregation kinetics is enhanced by increasing experimental temperatures (5-85 °C).27-29 Snapshots of Aβ(1-40) and Aβ(1-42) at different time scales at 400 K are extracted and illustrated in Figure 2. It can be seen that both peptides quickly move away from the initial nativelike conformation, which means that they unfold readily in the simulation timescales. A close look at Figure 2 reveals that: (i) the N-termini of Aβ(1-40) and Aβ(1-42) exhibit similar dynamic behaviors, namely, the residues 3-12 and 2022 are relatively flexible than other parts, which is in accordance with recent experimental reports;14,15 (ii) the dynamic behaviors of the C-termini of both peptides are distinct, that is, the C-terminus of Aβ(1-42) unfolds completely at about 1.5 ns; in contrast, a certain part of C-terminal helix in Aβ(1-40)
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Figure 3. Stereoviews of the hydrophobic interactions in the C-terminus of Aβ(1-40) and Aβ(1-42) at 1.5 ns of the MD simulations. The residues involved in the hydrophobic interactions are shown in Corey-Pauling-Koltun spheres. (a) Aβ(1-40); (b) Aβ(1-42). It can be seen that, in the C-terminus of Aβ(1-40), residues 39-40 (VV) form hydrophobic interactions with residues 34-36 (LMV), owing to the favorable spacial positions in R-helical conformation, which stabilizes the C-terminal helical conformation of Aβ(1-40). In comparison, the addition of the two residues 41-42 (IA) in Aβ(1-42) disrupts the hydrophobic core in Aβ(1-40). That is, residues 41-42 (IA) form relatively strong hydrophobic interactions with residues 39-40 (VV), which makes residues 34-36 (LMV) to close with residues 31-32 (II) to form a new hydrophobic core. Apparently, the double-centered hydrophobic core in Aβ(1-42) does not favor to stabilize the helix but rather behaves as a trigger to unravel the C-terminus of Aβ(1-42).
remains at 5 ns. Therefore, the C-terminus of Aβ(1-42) is more unfolded than that of Aβ(1-40), which is in good agreement with recent experimental findings.14,15 To explore the physical basis underlying the different dynamic behaviors of both Aβ peptides, we should consider the interaction patterns within the C-termini of both peptides. Since the C-termini of Aβ peptides contain no hydrophilic residues, hydrophobic interaction is likely to be the major player. In fact, hydrophobic interactions have proved to make great contributions to Aβ aggregation.30,31 Because of the favorable spacial positions in R-helical conformations between residues 39-40 (VV) and 34-36 (LMV), they form a hydrophobic core in the C-terminus of Aβ(1-40) that definitely stabilizes the peptide (Figure 3a and Figure 4). However, this hydrophobic core is destroyed in Aβ(1-42), because the additional residues 41-42 (IA) form hydrophobic interactions with residues 39-40 (VV), as revealed by the continuously strengthening of hydrophobic interactions (Figure 4). This induced the disruption of the hydrophobic interactions between residues 39-40 (VV) and residues 34-
36 (LMV), as reflected by the continuous weakening of corresponding hydrophobic interactions (Figure 4). As a result, residues 34-36 (LMV) tend to close with 31-32 (II) to form a new hydrophobic core (Figure 3b). Apparently, the doublecentered hydrophobic core in Aβ(1-42) does not favor to stabilize the helix but rather behaves as a trigger to unravel the C-terminus of Aβ(1-42). This provides a reasonable explanation for the distinct dynamic behavior of the C-terminus in Aβ(142) and Aβ(1-40). To support the above conclusion, MD simulations on a I41A mutant of Aβ(1-42) were also performed on the same level. The results indicate that the C-terminus of the I41A mutant of Aβ(1-42) still remains as R-helical conformation till the end of the simulation (Figure 2). As illustrated in Figure 1, the RMSD of the I41A mutant of Aβ(1-42) is relatively lower than that of Aβ(1-42) at 400 K, implying that the former possesses higher conformational stability comparing with the latter. Further observation revealed that the residues 39-40 (VV) in the I41A mutant of Aβ(1-42) formed a hydrophobic core with residues 34-36 (LMV) during the whole simulation, which is different
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Figure 4. Changes (average results of five parallel simulations) of hydrophobic interaction energies (estimated by “analyze” in the Discover module of InsightII program package) against the simulation time. (a) Energies between residues 39-40 (VV) and residues 34-36 (LMV) in the C-terminus of Aβ(1-40); (b) energies between residues 39-40 (VV) and residues 34-36 (LMV) in the C-terminus of Aβ(142); (c) energies between residues 39-40 (VV) and residues 41-42 (IA) in the C-terminus of Aβ(1-42); (d) energies between residues 39-40 (VV) and residues 34-36 (LMV) in the C-terminus of I41A mutant of Aβ(1-42). It can be seen that in the C-terminus of Aβ(140) the hydrophobic interactions change slightly during the simulation. In the C-terminus of Aβ(1-42), the hydrophobic interactions for residues 39-40 (VV) and residues 34-36 (LMV) weaken continuously, while those for residues 39-40 (VV) and residues 41-42 (IA) strengthen continuously. The changes of the hydrophobic interactions between residues 39-40 (VV) and residues 34-36 (LMV) of the I41A mutant of Aβ(1-42) against simulation time are different from those of Aβ(1-42) though similar to those of Aβ(1-40).
from the hydrophobic interactions pattern in Aβ(1-42), while consistent with that in Aβ(1-40) as mentioned above. This is also reflected by the fact that the changes of the hydrophobic interaction energies against simulation time of I41A mutant of Aβ(1-42) are different from those of Aβ(1-42) (parts d and b of Figure 4), while similar to those of Aβ(1-40) (parts d and a of Figure 4). In summary, based on the MD simulations of Aβ(1-40), Aβ(1-42) and I41A mutant of Aβ(1-42), it is revealed that the hydrophobic interactions play a crucial role in the C-terminus unfolding of Aβ(1-42), which results in the more unfolded C-terminus of Aβ(1-42) than that of Aβ(1-40). This finding provides some new clues to understanding the different aggregation and deposition propensities of both peptides and is helpful to elucidating the pathogenesis of AD as well. Acknowledgment. The authors thank the referees for their helpful suggestions. This work was supported by the National Basic Research Program of China (2003CB114400) and the National Natural Science Foundation of China (30570383 and 30700113).
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