D614G mutation in the SARS-CoV-2 spike protein enhances viral fitness by desensitizing it to temperature-dependent denaturation

Tzu-Jing Yang,Pei-Yu Yu,Yuan-Chih Chang,Shang-Te Danny Hsu

doi:10.1016/j.jbc.2021.101238

Abstract

The D614G mutation in the spike protein of SARS-CoV-2 alters the fitness of the virus, leading to the dominant form observed in the COVID-19 pandemic. However, the molecular basis of the mechanism by which this mutation enhances fitness is not clear. Here we demonstrated by cryo-electron microscopy that the D614G mutation resulted in increased propensity of multiple receptor-binding domains (RBDs) in an upward conformation poised for host receptor binding. Multiple substates within the one RBD-up or two RBD-up conformational space were determined. According to negative staining electron microscopy, differential scanning calorimetry, and differential scanning fluorimetry, the most significant impact of the mutation lies in its ability to eliminate the unusual cold-induced unfolding characteristics and to significantly increase the thermal stability under physiological pH. The D614G spike variant also exhibited exceptional long-term stability when stored at 37 °C for up to 2 months. Our findings shed light on how the D614G mutation enhances the infectivity of SARS-CoV-2 through a stabilizing mutation and suggest an approach for better design of spike protein-based conjugates for vaccine development.

Highlights

The COVID-19 pandemic is caused by the infection of SARS-CoV-2 [1]
We further conducted the negative staining electronic microscopy (NSEM) analysis coupled with differential scanning calorimetry (DSC) and differential scanning fluorimetry (DSF) to reveal that the D614G mutation eliminates the cold sensitivity of the original D614 and confers the resistance to the high temperature
Using cryo-EM single particle reconstruction aided by 3D variability analysis (3DVA) analysis [13, 14], we identified five distinct but populated clusters of conformations of S-D614G with varying degrees of receptor-binding domain (RBD)-up populations (Fig. 1, Fig. S1 and Table S1)

Summary

Results

Using cryo-EM single particle reconstruction aided by 3DVA analysis [13, 14], we identified five distinct but populated clusters of conformations of S-D614G with varying degrees of RBD-up populations (Fig. 1, Fig. S1 and Table S1). ACCELERATED COMMUNICATION: D614G reduced temperature sensitivity of SARS-CoV-2 spike

RBD-up θ

Discussion

Experimental procedures