A broad and potent neutralization epitope in SARS-related coronaviruses.

Many neutralizing antibodies (nAbs) elicited to ancestral severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) through natural infection and vaccination have reduced effectiveness to SARS-CoV-2 variants. Here, we show that therapeutic antibody ADG20 is able to neutralize SARS-CoV-2 variants of concern (VOCs) including Omicron (B.1.

A broad and potent neutralization epitope in SARS-related coronaviruses.

Many neutralizing antibodies (nAbs) elicited to ancestral SARS-CoV-2 through natural infection and vaccination generally have reduced effectiveness to SARS-CoV-2 variants. Here we show therapeutic antibody ADG20 is able to neutralize all SARS-CoV-2 variants of concern (VOCs) including Omicron (B.1.

SARS-CoV-2 Beta variant infection elicits potent lineage-specific and cross-reactive antibodies.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) Beta variant of concern (VOC) resists neutralization by major classes of antibodies from COVID-19 patients and vaccinated individuals. In this study, serum of Beta-infected patients revealed reduced cross-neutralization of wild-type virus. From these patients, we isolated Beta-specific and cross-reactive receptor-binding domain (RBD) antibodies.

An alternate conformation of HCV E2 neutralizing face as an additional vaccine target.

To achieve global elimination of hepatitis C virus (HCV), an effective cross-genotype vaccine is needed. The HCV envelope glycoprotein E2 is the main target for neutralizing antibodies (nAbs), which aid in HCV clearance and protection. E2 is structurally flexible and functions in engaging host receptors.

Structural basis of a shared antibody response to SARS-CoV-2.

Molecular understanding of neutralizing antibody responses to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) could accelerate vaccine design and drug discovery. We analyzed 294 anti-SARS-CoV-2 antibodies and found that immunoglobulin G heavy-chain variable region 3-53 (IGHV3-53) is the most frequently used IGHV gene for targeting the receptor-binding domain (RBD) of the spike protein. Co-crystal structures of two IGHV3-53-neutralizing antibodies with RBD, with or without Fab CR3022, at 2.

Structural basis of a public antibody response to SARS-CoV-2.

Molecular-level understanding of human neutralizing antibody responses to SARS-CoV-2 could accelerate vaccine design and facilitate drug discovery. We analyzed 294 SARS-CoV-2 antibodies and found that IGHV3-53 is the most frequently used IGHV gene for targeting the receptor binding domain (RBD) of the spike (S) protein. We determined crystal structures of two IGHV3-53 neutralizing antibodies +/- Fab CR3022 ranging from 2.

Genetic and structural insights into broad neutralization of hepatitis C virus by human V1-69 antibodies.

An effective vaccine to the antigenically diverse hepatitis C virus (HCV) must target conserved immune epitopes. Here, we investigate cross-neutralization of HCV genotypes by broadly neutralizing antibodies (bNAbs) encoded by the relatively abundant human gene family . We have deciphered the molecular requirements for cross-neutralization by this unique class of human antibodies from crystal structures of HCV E2 in complex with bNAbs.

Key gp120 Glycans Pose Roadblocks to the Rapid Development of VRC01-Class Antibodies in an HIV-1-Infected Chinese Donor.

VRC01-class antibodies neutralize diverse HIV-1 strains by targeting the conserved CD4-binding site. Despite extensive investigations, crucial events in the early stage of VRC01 development remain elusive. We demonstrated how VRC01-class antibodies emerged in a Chinese donor by antigen-specific single B cell sorting, structural and functional studies, and longitudinal antibody and virus repertoire analyses.

Affinity Maturation of a Potent Family of HIV Antibodies Is Primarily Focused on Accommodating or Avoiding Glycans.

The high-mannose patch on the HIV-1 envelope (Env) glycoprotein is the epicenter for binding of the potent broadly neutralizing PGT121 family of antibodies, but strategies for generating such antibodies by vaccination have not been defined. We generated structures of inferred antibody intermediates by X-ray crystallography and electron microscopy to elucidate the molecular events that occurred during evolution of this family. Binding analyses revealed that affinity maturation was primarily focused on avoiding, accommodating, or binding the N137 glycan.

Complete epitopes for vaccine design derived from a crystal structure of the broadly neutralizing antibodies PGT128 and 8ANC195 in complex with an HIV-1 Env trimer.

The HIV-1 envelope gp160 glycoprotein (Env) is a trimer of gp120 and gp41 heterodimers that mediates cell entry and is the primary target of the humoral immune response. Broadly neutralizing antibodies (bNAbs) to HIV-1 have revealed multiple epitopes or sites of vulnerability, but mapping of most of these sites is incomplete owing to a paucity of structural information on the full epitope in the context of the Env trimer. Here, a crystal structure of the soluble BG505 SOSIP gp140 trimer at 4.

Design and structure of two HIV-1 clade C SOSIP.664 trimers that increase the arsenal of native-like Env immunogens.

A key challenge in the quest toward an HIV-1 vaccine is design of immunogens that can generate a broadly neutralizing antibody (bnAb) response against the enormous sequence diversity of the HIV-1 envelope glycoprotein (Env). We previously demonstrated that a recombinant, soluble, fully cleaved SOSIP.664 trimer based on the clade A BG505 sequence is a faithful antigenic and structural mimic of the native trimer in its prefusion conformation.

Recombinant HIV envelope trimer selects for quaternary-dependent antibodies targeting the trimer apex.

Broadly neutralizing antibodies (bnAbs) targeting the trimer apex of HIV envelope are favored candidates for vaccine design and immunotherapy because of their great neutralization breadth and potency. However, methods of isolating bnAbs against this site have been limited by the quaternary nature of the epitope region. Here we report the use of a recombinant HIV envelope trimer, BG505 SOSIP.

Structural evolution of glycan recognition by a family of potent HIV antibodies.

The HIV envelope glycoprotein (Env) is densely covered with self-glycans that should help shield it from recognition by the human immune system. Here, we examine how a particularly potent family of broadly neutralizing antibodies (Abs) has evolved common and distinct structural features to counter the glycan shield and interact with both glycan and protein components of HIV Env. The inferred germline antibody already harbors potential binding pockets for a glycan and a short protein segment.

Hepatitis C virus E2 envelope glycoprotein core structure.

Hepatitis C virus (HCV), a Hepacivirus, is a major cause of viral hepatitis, liver cirrhosis, and hepatocellular carcinoma. HCV envelope glycoproteins E1 and E2 mediate fusion and entry into host cells and are the primary targets of the humoral immune response. The crystal structure of the E2 core bound to broadly neutralizing antibody AR3C at 2.

Supersite of immune vulnerability on the glycosylated face of HIV-1 envelope glycoprotein gp120.

A substantial proportion of the broadly neutralizing antibodies (bnAbs) identified in certain HIV-infected donors recognize glycan-dependent epitopes on HIV-1 gp120. Here we elucidate how the bnAb PGT 135 binds its Asn332 glycan-dependent epitope from its 3.1-Å crystal structure with gp120, CD4 and Fab 17b.

Disulfide bond stabilization of the hexameric capsomer of human immunodeficiency virus.

The human immunodeficiency virus type 1 capsid is modeled as a fullerene cone that is composed of approximately 250 hexamers and 12 pentamers of the viral CA protein. Structures of CA hexamers have been difficult to obtain because the hexamer-stabilizing interactions are inherently weak, and CA tends to spontaneously assemble into capsid-like particles. Here, we describe a two-step biochemical strategy to obtain soluble CA hexamers for crystallization.

X-ray structures of the hexameric building block of the HIV capsid.

The mature capsids of HIV and other retroviruses organize and package the viral genome and its associated enzymes for delivery into host cells. The HIV capsid is a fullerene cone: a variably curved, closed shell composed of approximately 250 hexamers and exactly 12 pentamers of the viral CA protein. We devised methods for isolating soluble, assembly-competent CA hexamers and derived four crystallographically independent models that define the structure of this capsid assembly unit at atomic resolution.

Profiling of membrane protein variants in a baculovirus system by coupling cell-surface detection with small-scale parallel expression.

Production of structure-grade mammalian membrane proteins in substantial quantities has been hindered by a lack of methods for effectively profiling multiple constructs expression in higher eukaryotic systems such as insect or mammalian cells. To address this problem, a specialized small-scale eukaryotic expression platform by Thomson Instrument Company (Vertiga-IM) was developed and used in tandem with a Guava EasyCyte microcapillary 96-well cytometer to monitor cell density and health and evaluate membrane protein expression. Two proof of concept experiments were conducted using the human beta(2)-adrenergic receptor (beta(2)AR) and the gap junction protein connexin26 (Cx26) in a baculovirus expression system.

Cloning and expression of multiple integral membrane proteins from Mycobacterium tuberculosis in Escherichia coli.

Seventy integral membrane proteins from the Mycobacterium tuberculosis genome have been cloned and expressed in Escherichia coli. A combination of T7 promoter-based vectors with hexa-His affinity tags and BL21 E. coli strains with additional tRNA genes to supplement sparsely used E.