Membrane fusion and immune evasion by the spike protein of SARS-CoV-2 Delta variant

Abstract

The Delta variant of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has outcompeted previously prevalent variants and become a dominant strain worldwide. We report the structure, function, and antigenicity of its full-length spike (S) trimer as well as those of the Gamma and Kappa variants, and compare their characteristics with the G614, Alpha, and Beta variants. Delta S can fuse membranes more efficiently at low levels of cellular receptor angiotensin converting enzyme 2 (ACE2), and its pseudotyped viruses infect target cells substantially faster than the other five variants, possibly accounting for its heightened transmissibility. Each variant shows different rearrangement of the antigenic surface of the amino-terminal domain of the S protein but only makes produces changes in the receptor binding domain (RBD), making the RBD a better target for therapeutic antibodies.

Fig. 1.. More efficient membrane fusion by the Delta variant than other variants.

(A) Time course of cell-cell fusion mediated by various full-length S proteins, as indicated by the labels, with HEK293 cells with no exogenous ACE2. (B) Cell-cell fusion mediated by various full-length S proteins with HEK293 cells transfected with low levels (0 to 0.25 ng) of ACE2 expression constructs. (C) Time course of infection HEK293-ACE2 cells by MLV-based, pseudotyped viruses by various SARS-CoV-2 variant S constructs containing a CT deletion in a single cycle. Infection was initiated by mixing viruses and target cells, and viruses were washed out at each time point as indicated. The full time course and concentration series are shown in fig. S3. The experiments were repeated at least three times, with independent samples each giving similar results.

Fig. 2.. Antigenic properties of purified full-length SARS-CoV-2 S proteins.

Biolayer interferometry (BLI) analysis of the association of prefusion S trimers derived from the G614 “parent” strain (B.1) and the Gamma (B.1.1.28), Kappa (B.1.617.1), and Delta (B.1.617.2) variants with soluble ACE2 constructs and with a panel of antibodies representing five epitopic regions on the RBD and NTD (see fig. S4A) (32). For ACE2 binding, purified S proteins were immobilized to AR2G biosensors and dipped into wells containing ACE2 at various concentrations. For antibody binding, various antibodies were immobilized to AHC biosensors and dipped into wells containing each purified S protein at different concentrations. Binding kinetics were evaluated by a 1:1 Langmuir model except for dimeric ACE2 and antibody G32B6 targeting the RBD-2, which were analyzed by a bivalent binding model. All K_D values for multivalent interactions with antibody IgG or dimeric ACE2 and trimeric S protein are the apparent affinities with avidity effects. Sensorgrams are in black and fits are in red. Binding constants highlighted by underlines were estimated by steady state analysis as described in the materials and methods. RU, response unit. Binding constants are summarized both here and in table S1. All experiments were repeated at least twice with essentially identical results.

Fig. 3.. Cryo-EM structures of full-length SARS-CoV-2 S proteins from the Delta, Kappa, and Gamma variants.

(A to C) The structures of the closed prefusion conformation and two one-RBD-up conformations of the Delta S trimer are shown in the ribbon diagrams, with one protomer colored as follows: NTD in blue, RBD in cyan, CTD1 in green, CTD2 in light green, S2 in light blue, the 630 loop in red, and the FPPR in magenta. (D to F) The structures of the closed prefusion conformation and two one-RBD-up conformation of the Kappa S trimer are shown in the ribbon diagrams, with the same color scheme as in (A). (G) and (H) The structures of the two one-RBD-up conformations of the Gamma S trimer are shown in the ribbon diagrams with the same color scheme as in (A). All mutations in the three variants, as compared to the original virus (D614), are highlighted in the sphere model. (I) Structures in the Delta closed trimer of segments (residues 617 to 644) containing the 630 loop (red) and segments (residues 823 to 862) containing the FPPR (magenta) from each of the three protomers (A), (B), and (C). The position of each RBD is indicated. Dashed lines indicate gaps in the chain trace (disordered loops).

Fig. 4.. Structural impact of mutations in Delta S.

(A) Superposition of the NTD structure of the Delta S trimer (blue) with the NTD of the G614 S trimer (PDB ID: 7KRQ) (yellow). Locations of mutations T19R, G142D, E156G, and deletion of F157 and R158 are indicated; these residues are shown in the stick model. The N-terminal segment, as well as loops 143 to 154 and 173 to 187, are rearranged between the two structures and highlighted in darker colors. (B) Top view of panel (A). (C) Superposition of the RBD structure of the Delta S trimer (cyan) with the RBD of the G614 S trimer (yellow). Locations of mutations L452R and T478K are indicated; these residues are shown in the stick model. (D) A close-up view of superposition of the Delta S2 (light blue) with the S2 of the G614 S trimer (yellow) near residue 950. Locations of the D950N mutation and charged residues in the vicinity including Lys⁹⁴⁷, Arg¹⁰¹⁴, and Glu¹⁰¹⁷ from protomer A and Glu⁷⁷³, Lys⁷⁷⁶, Glu⁷⁸⁰, and Arg¹⁰¹⁹ from the protomer B are indicated. All aforementioned residues are shown in the stick model.

Fig. 5.. Structural impact of mutations in the Kappa and Gamma S proteins.

(A) Superposition of the NTD structure of the Kappa S trimer (blue) with the NTD of the G614 S trimer (yellow). Locations of mutations E154K and Q218H, as well as Arg¹⁰² which forms a salt bridge with Glu¹⁵⁴ in the G614 structure, are indicated; these residues are shown in the stick model. The 173 to 187 loop in the G614 trimer is highlighted in a darker color; it becomes disordered in the Kappa trimer. (B) Superposition of the RBD structure of the Kappa S trimer (cyan) with the RBD of the G614 S trimer (yellow). Locations of mutations L452R and E484Q are indicated; these residues are shown in the stick model. (C) A view of superposition of the NTD structures of the Gamma (blue) and G614 (yellow; PDB ID: 7KRR) S trimers in the one-RBD-up conformation. Locations of mutations L18F, T20N, P26S, D138Y, and R190S are indicated, as well as the N-linked glycan attached to Asn²⁰ in the Gamma structure; these residues are shown in the stick model. (D) Superposition of the RBD structure of the Gamma S trimer (cyan) with the RBD of the G614 S trimer (yellow). Locations of mutations K417T, E484K, and N501Y are indicated, and these residues are shown in the stick model.