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Dynamically-driven inactivation of the catalytic machinery of the SARS 3C-like protease by the N214A mutation on the extra domain.

Shi J, Han N, Lim L, Lua S, Sivaraman J, Wang L, Mu Y, Song J - PLoS Comput. Biol. (2011)

Bottom Line: Therefore, the N214A mutation appears to trigger the dramatic change of the enzyme dynamics in the context of the dimeric form which ultimately inactivates the catalytic machinery.The present MD simulations represent the longest reported so far for the SARS-CoV 3CLpro, unveiling that its catalysis is critically dependent on the dynamics, which can be amazingly modulated by the extra domain.Consequently, mediating the dynamics may offer a potential avenue to inhibit the SARS-CoV 3CLpro.

View Article: PubMed Central - PubMed

Affiliation: Department of Biological Sciences, Faculty of Science, National University of Singapore, Singapore.

ABSTRACT
Despite utilizing the same chymotrypsin fold to host the catalytic machinery, coronavirus 3C-like proteases (3CLpro) noticeably differ from picornavirus 3C proteases in acquiring an extra helical domain in evolution. Previously, the extra domain was demonstrated to regulate the catalysis of the SARS-CoV 3CLpro by controlling its dimerization. Here, we studied N214A, another mutant with only a doubled dissociation constant but significantly abolished activity. Unexpectedly, N214A still adopts the dimeric structure almost identical to that of the wild-type (WT) enzyme. Thus, we conducted 30-ns molecular dynamics (MD) simulations for N214A, WT, and R298A which we previously characterized to be a monomer with the collapsed catalytic machinery. Remarkably, three proteases display distinctive dynamical behaviors. While in WT, the catalytic machinery stably retains in the activated state; in R298A it remains largely collapsed in the inactivated state, thus implying that two states are not only structurally very distinguishable but also dynamically well separated. Surprisingly, in N214A the catalytic dyad becomes dynamically unstable and many residues constituting the catalytic machinery jump to sample the conformations highly resembling those of R298A. Therefore, the N214A mutation appears to trigger the dramatic change of the enzyme dynamics in the context of the dimeric form which ultimately inactivates the catalytic machinery. The present MD simulations represent the longest reported so far for the SARS-CoV 3CLpro, unveiling that its catalysis is critically dependent on the dynamics, which can be amazingly modulated by the extra domain. Consequently, mediating the dynamics may offer a potential avenue to inhibit the SARS-CoV 3CLpro.

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Dynamic behavior of the Phe140-His172 interaction.Time-trajectories of the centroid distances between the aromatic rings of Phe140 and His172 of WT (a–c); R298A (d–f) and N214A (g–i) in three independent simulations. Time-trajectories of the Chi1 dihedral angle of His172 of WT (j–l); R298A (m–o) and N214A (p–r). Time-trajectories of the Chi2 dihedral angle of His172 of WT (s–u); R298A (v–x) and N214A (y–aa).
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pcbi-1001084-g008: Dynamic behavior of the Phe140-His172 interaction.Time-trajectories of the centroid distances between the aromatic rings of Phe140 and His172 of WT (a–c); R298A (d–f) and N214A (g–i) in three independent simulations. Time-trajectories of the Chi1 dihedral angle of His172 of WT (j–l); R298A (m–o) and N214A (p–r). Time-trajectories of the Chi2 dihedral angle of His172 of WT (s–u); R298A (v–x) and N214A (y–aa).

Mentions: Figure 8 shows the dynamical behaviors for the aromatic interaction between Phe140 and His172. In WT, the centroid distance remains short and dynamically stable in three simulations, with average values of 5.16 and 5.12 Å respectively for two protomers. By contrast, in R298A, this distance remains large as observed in the crystal structure, with an average value of 8.15 Å, indicating the total absence of this stacking interaction in the whole 30 ns simulations. For N214A, the dynamic behavior of this distance is very similar to that for WT, despite having slightly larger average values of 5.42 and 5.14 Å respectively for two protomers. As for the side chain conformations, the His172 Chi1 angles have both a very similar value as well as similar dynamical behavior in all three enzymes except that in one simulation of N214A, one protomer jumps to sample another conformation (Figure 8r). However, significant dynamics are observed for the N214A Chi2, which jumps to sample several conformational clusters in simulations (Figures 8y–8aa).


Dynamically-driven inactivation of the catalytic machinery of the SARS 3C-like protease by the N214A mutation on the extra domain.

Shi J, Han N, Lim L, Lua S, Sivaraman J, Wang L, Mu Y, Song J - PLoS Comput. Biol. (2011)

Dynamic behavior of the Phe140-His172 interaction.Time-trajectories of the centroid distances between the aromatic rings of Phe140 and His172 of WT (a–c); R298A (d–f) and N214A (g–i) in three independent simulations. Time-trajectories of the Chi1 dihedral angle of His172 of WT (j–l); R298A (m–o) and N214A (p–r). Time-trajectories of the Chi2 dihedral angle of His172 of WT (s–u); R298A (v–x) and N214A (y–aa).
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3044768&req=5

pcbi-1001084-g008: Dynamic behavior of the Phe140-His172 interaction.Time-trajectories of the centroid distances between the aromatic rings of Phe140 and His172 of WT (a–c); R298A (d–f) and N214A (g–i) in three independent simulations. Time-trajectories of the Chi1 dihedral angle of His172 of WT (j–l); R298A (m–o) and N214A (p–r). Time-trajectories of the Chi2 dihedral angle of His172 of WT (s–u); R298A (v–x) and N214A (y–aa).
Mentions: Figure 8 shows the dynamical behaviors for the aromatic interaction between Phe140 and His172. In WT, the centroid distance remains short and dynamically stable in three simulations, with average values of 5.16 and 5.12 Å respectively for two protomers. By contrast, in R298A, this distance remains large as observed in the crystal structure, with an average value of 8.15 Å, indicating the total absence of this stacking interaction in the whole 30 ns simulations. For N214A, the dynamic behavior of this distance is very similar to that for WT, despite having slightly larger average values of 5.42 and 5.14 Å respectively for two protomers. As for the side chain conformations, the His172 Chi1 angles have both a very similar value as well as similar dynamical behavior in all three enzymes except that in one simulation of N214A, one protomer jumps to sample another conformation (Figure 8r). However, significant dynamics are observed for the N214A Chi2, which jumps to sample several conformational clusters in simulations (Figures 8y–8aa).

Bottom Line: Therefore, the N214A mutation appears to trigger the dramatic change of the enzyme dynamics in the context of the dimeric form which ultimately inactivates the catalytic machinery.The present MD simulations represent the longest reported so far for the SARS-CoV 3CLpro, unveiling that its catalysis is critically dependent on the dynamics, which can be amazingly modulated by the extra domain.Consequently, mediating the dynamics may offer a potential avenue to inhibit the SARS-CoV 3CLpro.

View Article: PubMed Central - PubMed

Affiliation: Department of Biological Sciences, Faculty of Science, National University of Singapore, Singapore.

ABSTRACT
Despite utilizing the same chymotrypsin fold to host the catalytic machinery, coronavirus 3C-like proteases (3CLpro) noticeably differ from picornavirus 3C proteases in acquiring an extra helical domain in evolution. Previously, the extra domain was demonstrated to regulate the catalysis of the SARS-CoV 3CLpro by controlling its dimerization. Here, we studied N214A, another mutant with only a doubled dissociation constant but significantly abolished activity. Unexpectedly, N214A still adopts the dimeric structure almost identical to that of the wild-type (WT) enzyme. Thus, we conducted 30-ns molecular dynamics (MD) simulations for N214A, WT, and R298A which we previously characterized to be a monomer with the collapsed catalytic machinery. Remarkably, three proteases display distinctive dynamical behaviors. While in WT, the catalytic machinery stably retains in the activated state; in R298A it remains largely collapsed in the inactivated state, thus implying that two states are not only structurally very distinguishable but also dynamically well separated. Surprisingly, in N214A the catalytic dyad becomes dynamically unstable and many residues constituting the catalytic machinery jump to sample the conformations highly resembling those of R298A. Therefore, the N214A mutation appears to trigger the dramatic change of the enzyme dynamics in the context of the dimeric form which ultimately inactivates the catalytic machinery. The present MD simulations represent the longest reported so far for the SARS-CoV 3CLpro, unveiling that its catalysis is critically dependent on the dynamics, which can be amazingly modulated by the extra domain. Consequently, mediating the dynamics may offer a potential avenue to inhibit the SARS-CoV 3CLpro.

Show MeSH
Related in: MedlinePlus