Experimental Evidence for Enhanced Receptor Binding by Rapidly Spreading SARS-CoV-2 Variants

Abstract

Rapidly spreading new variants of SARS-CoV-2 carry multiple mutations in the viral spike protein which attaches to the angiotensin converting enzyme 2 (ACE2) receptor on host cells. Among these mutations are amino acid changes N501Y (lineage B.1.1.7, first identified in the UK), and the combination N501Y, E484K, K417N (B.1.351, first identified in South Africa), all located at the interface on the receptor binding domain (RBD). We experimentally establish that RBD containing the N501Y mutation results in 7-fold stronger binding to the hACE2 receptor than wild type RBD. The E484K mutation only slightly enhances the affinity for the receptor, while K417N attenuates affinity. As a result, RBD from B.1.351 containing all three mutations binds 3-fold stronger to hACE2 than wild type RBD but 2-fold weaker than N501Y. However, the recently emerging double mutant E484K/N501Y binds even stronger than N501Y. The independent evolution of lineages containing mutations with different effects on receptor binding affinity, viral transmission and immune evasion underscores the importance of global viral genome surveillance and functional characterization.

Keywords: angiotensin converting enzyme 2 receptor; coronavirus; mutations; spike receptor binding domain; surface plasmon resonance.

Graphical abstract

Figure 1

Effects of amino acid mutations on molecular interactions and strength of the interface between hACE2 and the SARS-CoV-2 receptor binding domain. (A) Location of residues N501, E484 and K417 indicated by blue spheres at the interface of SARS-CoV-2 receptor binding domain (white cartoon representation) and the hACE2 ectodomain (orange surface representation) in 6m0j.pdb (24). (B) Details of the interactions with RBD N501 forming a hydrogen bond with hACE2 Y41, RBD E484 forming an ion-pair with hACE2 K31, and RBD K417 forming a salt-bridge (ion-pair plus hydrogen bond) with hACE2 D30. C) Amino acid changes in RBDs of different SARS-CoV-2 lineages. D) Cartoon visualizing SPR setup using Biacore T100. (E) Sensorgrams of increasing concentrations of RBD binding to immobilized hACE ectodomain (colored datapoints) with fit of a 1:1 binding model (thin black lines) for wild type and mutant RBDs. (F) Affinity of RBD mutants relative to wild type RBD.