In addition, the side chain of T345 in the RBD hydrogen bonds with the main chain carbonyl of K92 in the LC CDR3 ( Fig. The other major interaction site involves RBD residue N440, which engages in a direct hydrogen bond with W33 from CDR1 and aligns parallel to the side chain of Y52 in the CDR2 to make CH-π hydrophobic interaction in the HC ( Fig. The T102 hydroxyl in the CDR3 HC also hydrogen bonds with the RBD N448 side chain ( Fig. 2C) and has the potential for a cation-π stacking interaction involving Y32 from LC CDR1. Specifically, the guanidine group of R346 engages in multiple hydrogen bonds involving T102 and Y91 from the HC and LC CDR3, respectively ( Fig. RBD residue R346 is the main contact point and is sandwiched between the HC CDR3 and LC CDR1 and CDR3 regions. Most of the interactions are mediated through the HC and LC CDR3 regions, and the epitope aligns with RBD-5/class 3 antibodies ( 12, 13). The interaction buried a total surface area of ~737 Å 2 with HC and LC contributing ~60 and ~40% of the total interaction, respectively ( Fig. The cryo-EM structure showed that 002-S21F2 binds to the outer face of the RBD that is accessible in both “down” and “up” conformations and is outside the ACE2-binding motif ( Fig. To define the molecular features conferring epitope recognition and to understand the mechanism of the broad neutralization spectrum of 002-S21F2 against SARS-CoV-2 variants, we determined the cryo-EM structures of 002-S21F2 full-length IgG in complex with WA.1 and Omicron spike-6P at 3.7 and 4.1 Å, respectively ( Fig. Antibody 002-S21F2 was the most potent among all the mAbs that neutralized live SARS-CoV-2 WA.1 and thus was selected for comprehensive downstream characterization. S2A), and 18 (37.5%) successfully neutralized live virus with 50% inhibitory concentration (IC 50) values ranging from 0.05 to 17 μg/ml (fig. Of these mAbs, 48 blocked the ACE2-RBD interaction (fig. Most of these mAbs had a low frequency of somatic hypermutations (SHM) in both their HC and LC, suggesting that they were recently recruited from a naive B cell pool which is typical of a primary infection (fig. S1E) ( 16), as well as enriched usage of HC and LC variable region genes belonging to the IGHV3, IGKV1, and IGLV1 families (fig.
These mAbs showed an average CDR3 (complementarity-determining region 3) length of 16.3 amino acids, which is typical of a human immunoglobulin G (IgG) repertoire (fig. RBD-based enzyme-linked immunosorbent assay (ELISA) screening resulted in the identification of 92 SARS-CoV-2–specific mAbs (fig. In total, we sorted 804 SARS-CoV-2 RBD fluorescent probe binding class-switched B cells, amplified 398 (~50%) paired heavy- (HC) and light-chain (LC) antibody gene sequences, and successfully cloned and expressed 208 antibodies ( Fig. S1, A to C) for the generation of SARS-CoV-2 RBD-specific mAbs. We selected five individuals (table S1) who had high SARS-CoV-2 RBD binding titers, high neutralization titers to live SARS-CoV-2 WA.1, and had detectable frequencies of RBD-specific memory B cells (fig.
We previously evaluated the humoral immune responses in 42 COVID-19–recovered individuals who had experienced mild symptoms after the ancestral Wuhan strain (WA.1) transmission in the year 2020 ( 15). The discovery of 002-S21F2 and the broadly neutralizing epitope it targets have timely implications for developing a broad range of therapeutic and vaccine interventions against SARS-CoV-2 variants including Omicron sublineages. Structural studies of 002-S21F2 in complex with spike trimers of Omicron and WA.1 showed that it targets a conformationally conserved epitope on the outer face of RBD (class 3 surface) outside the ACE2-binding motif, thereby providing a mechanistic insights for its broad neutralization activity. In this study, by characterizing several human monoclonal antibodies (mAbs) isolated from single B cells of the COVID-19–recovered individuals in India who experienced ancestral Wuhan strain (WA.1) of SARS-CoV-2 during early stages of the pandemic, we found a receptor binding domain (RBD)–specific mAb 002-S21F2 that has rare gene usage and potently neutralized live viral isolates of SARS-CoV-2 variants including Alpha, Beta, Gamma, Delta, and Omicron sublineages (BA.1, BA.2, BA.2.12.1, BA.4, and BA.5) with IC 50 ranging from 0.02 to 0.13 μg/ml. Suthar, Rafi Ahmed, Eric Ortlund, Amit Sharma, Kaja Murali-Krishna, and Anmol Chandele Show Fewer Davis, Jens Wrammert, Sucheta Godbole, Amy R. Roback, Grace Mantus, Pawan Kumar Goel, Manju Rahi, Carl W. Davis-Gardner, Venkata Viswanadh Edara, Susanne Linderman, Kaustuv Nayak, Kritika Dixit, Pragati Sharma, Prashant Bajpai, Vanshika Singh, Filipp Frank, Narayanaiah Cheedarla, Hans P. Sanjeev Kumar, Anamika Patel, Lilin Lai, Chennareddy Chakravarthy, Rajesh Valanparambil, Elluri Seetharami Reddy, Kamalvishnu Gottimukkala, … Show All …, Meredith E.
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