A pH-dependent switch mediates conformational masking of SARS-CoV-2 spike

Tongqing Zhou, Yaroslav Tsybovsky, Adam S. Olia, Jason Gorman, Micah A. Rapp, Gabriele Cerutti, Phinikoula S. Katsamba, Alexandra Nazzari, Arne Schon, Pengfei D. Wang, Jude Bimela, Wei Shi, I-Ting Teng, Baoshan Zhang, Jeffrey C. Boyington, Gwo-Yu Chuang, Jared M. Sampson, Mallika Sastry, Tyler Stephens, Jonathan Stuckey, Shuishu Wang, Richard A. Friesner, David D. Ho, John R. Mascola, Lawrence Shapiro, Peter D. Kwong

SARS-CoV-2 has emerged as a global pathogen, sparking urgent vaccine development efforts with the trimeric spike. However, the inability of antibodies like CR3022, which binds a cryptic spike epitope with nanomolar affinity, to neutralize virus, suggests a spike-based means of neutralization escape. Here, we show the SARS-CoV-2 spike to have 10% the unfolding enthalpy of a globular protein at physiological pH, where it is recognized by antibodies like CR3022, and up to 10-times more unfolding enthalpy at endosomal pH, where it sheds such antibodies, suggesting that the spike evades potentially neutralizing antibody through a pH-dependent mechanism of conformational masking. To understand the compatibility of this mechanism with ACE2-receptor interactions, we carried out binding measurements and determined cryo-EM structures of the spike recognizing up to three ACE2 molecules at both physiological and endosomal pH. In the absence of ACE2, cryo-EM analyses indicated lower pH to reduce conformational heterogeneity. Single-receptor binding domain (RBD)-up conformations dominated at pH 5.5, resolving into a locked all-down conformation at lower pH through lowering of RBD and refolding of a pH-dependent switch. Notably, the emerging Asp614Gly strain partially destabilizes the switch that locks RBD down, thereby enhancing functional interactions with ACE2 while reducing evasion by conformational masking.

 doi: https://doi.org/10.1101/2020.07.04.187989

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