In a recent Current Research in Structural Biology study, researchers demonstrate that the energy landscape of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) exhibits significant conformational variation.
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Study: Energy landscape of the SARS-CoV-2 reveals extensive conformational heterogeneity. Image Credit: CROCOTHERY / Shutterstock.com
Background
Cryo-electron microscopy (cryo-EM) has been used to develop several structural paradigms of the SARS-CoV-2 spike (S) protein. However, these models and their accompanying electrostatic potential maps depict an unknown combination of conformations arising from the basal energy landscape of the S protein.
Similar to other proteins, certain conformational motions of SARS-CoV-2 Ss are assumed to be biophysically important. Yet, they cannot be explained just by static models.
About the study
In the current study, researchers present the energy profile of the glycosylated SARS-CoV-2 S protein using experimental cryo-EM images. The team also discovered how the glycan barrier was responsible for the stability of the observed low-energy conformations.
The researchers assessed the energy landscape of the S protein in a restricted space generated from experimental cryo-EM snapshots. Geometric machine-learning (ManifoldEM) was used to derive the spectrum of conformational motions from cryo-EM images.
The researchers then rebuilt the three-dimensional (3D) density of low-energy conformations on this landscape in the region of up-state. Density information was then interpreted in all-atoms detail using molecular dynamics (MD) simulations. Finally, the authors presented the local and global conformations of the S protein with an emphasis on the conformational variability of key binding sites.
Study findings
A wide range of iso-energetic conformations linked with the interacting sites of the receptor-binding domain (RBD) within the SARS-CoV-2 S protein that had not been previously identified. The conformational variability found in the current modeling, including 400 nanoseconds (ns) of string simulations and 32 ns of molecular dynamics flexible fitting (MDFF) simulations, was significantly higher than what can be achieved with 10 milliseconds (ms) of brute-force MD.
The experimentally observed horseshoe-shaped energy profile demonstrates that the simple rigid one-up RBD was insufficient to represent the complicated multi-dimensional conformational movements of the SARS-CoV-2 S protein.
The coordinated movements of the RBD have regional impacts that result in significant conformational variability that is relative to the holo or apo structures separately. Almost all of the binding pockets investigated here demonstrated evidence of a conformational selection process, in which particular minimum-energy sites were more or less favorable for binding.
Of the six analyzed binding sites, five, including those interacting with angiotensin-converting enzyme 2 (ACE2) receptor, two distinct antibodies, linoleic acid (LA), and mini-proteins exhibited altered conformations among their established apo- and holo-conformations. This was also reported during the one RBD-up global S conformation.
The apo-to-holo transition was evocative of a conformational preequilibrium. Only the AB-C135's binding site remained in the apo-state in all of the free energy-minimizing models tested, thereby indicating the need for an induced-fit mechanism for the docking of this antibody to the S.
In locations 51-59 in the LA binding pocket of the up RBD, the N234 glycan on chain A forms hydrogen bonds with residues 388 (Asn) and 387 (Leu). This interaction keeps the binding site open and results in super-open states relative to the holo 6ZB4/6ZB5 and apo 6VXX RBD structures.
This event explains the stability and instability of these low conformations associated with glycan shields. The N165 glycan exhibited similar pairwise potential energies for interactions with counterclockwise chain RBD.
Regions with solvent accessible surface area (SASA) values greater than those found in 6VSB were thought to be more prone to binding, which might have therapeutic effects. Thus, considering the vast conformational flexibility of key binding sites, incorporating a wide variety of low-energy conformations could potentially give rise to novel drug development approaches.
Conclusions
The study findings illustrate the energy landscape of the SARS-CoV-2 glycosylated S protein. Furthermore, the researchers revealed the diversity of low-energy conformations in the region of its open state, including the one RBD-upstate, using experimental cryo-EM images. The obtained atomic refinement exposed local and global molecular translocations that were not visible in a typical one RBD-up cryo-EM model.
The current study also demonstrated a multitude of openness in global conformations of the one RBD-upstate that were not disclosed by single-model analyses of the density maps. Further, it revealed conformations that coincided with the reported models.
Overall, the study highlights the flexible character of biological macromolecules like the SARS-CoV-2 RBD by looking beyond the static structures adapted to heavily averaged maps, which might have consequences for new medications. This unified modeling unveils RBD binding site conformations that were not evident in previously published apo-up structures or microsecond-long MD simulations.
Taken together, the present study provides a methodology for future research on describing the most likely up to down transitions of the SARS-CoV-2 RBD by reprocessing available cryoEM data.
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