Fine epitope mapping of monoclonal antibodies against hemagglutinin of a highly pathogenic H5N1 influenza virus using yeast surface display

Han, Thomas; Sui, Jianhua; Bennett, A. J.; Liddington, Robert; Donis, Ruben O.; Zhu, Quan; Marasco, Wayne A.

doi:10.1016/j.bbrc.2011.04.139

Cited by 31 publications

(19 citation statements)

References 12 publications

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“…1C). These key H5 residues do not overlap any known epitopes described above or detected with other human and mouse antibodies (38)(39)(40)(41)(42)(43)(44)(45)(46)(47)(48)(49)(50)(51). The four H5M9 epitope key residues do not overlap known H1 antigenic sites (36), but positions 53, 274, and 276 correspond to site C of H3 viruses (35).…”

Section: Discussionmentioning

confidence: 62%

A Unique and Conserved Neutralization Epitope in H5N1 Influenza Viruses Identified by an Antibody against the A/Goose/Guangdong/1/96 Hemagglutinin

Zhu

Guo

Jiang

et al. 2013

J Virol

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Section: Discussionmentioning

confidence: 62%

A Unique and Conserved Neutralization Epitope in H5N1 Influenza Viruses Identified by an Antibody against the A/Goose/Guangdong/1/96 Hemagglutinin

Zhu

Guo

Jiang

et al. 2013

J Virol

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“…In addition, the 65C6 epitope also partially overlaps epitopes detected by some human MAb, i.e., FLA5.10 at P118 (P122 according to H3 numbering), FLD21.140 at S121, Y164, and T167 (S125, Y168, and T171 according to H3 numbering) (8), AVFLuigG01 at P118, Y164, and T167 (P122, Y168, and T171 according to H3 numbering) (9) (Fig. 1H), and mouse MAb NR2728 at S121 (S125 according to H3 numbering) (10) (Fig. 1I).…”

mentioning

confidence: 99%

Unraveling of a Neutralization Mechanism by Two Human Antibodies against Conserved Epitopes in the Globular Head of H5 Hemagglutinin

Qian

Zuo

et al. 2013

J Virol

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The rapid spread of highly pathogenic avian influenza (HPAI) H5N1 virus underscores the importance of effective antiviral treatment. Previously, we developed human monoclonal antibodies 65C6 and 100F4 that neutralize almost all (sub)clades of HPAI H5N1. The conserved 65C6 epitope was mapped to the globular head of HA. However, neither the 100F4 epitope nor the neutralization mechanism by these antibodies was known. In this study, we determined the 100F4 epitope and unraveled a neutralization mechanism by antibodies 65C6 and 100F4. Influenza A virus infection is a persistent threat to public health worldwide. High-affinity neutralizing antibodies against conserved epitopes could provide immunity to diverse influenza virus strains and protection against future pandemic viruses. Previously, we developed human monoclonal antibodies (MAb) 65C6 and 100F4 that potently neutralize all H5 clades and subclades of highly pathogenic avian influenza (HPAI) H5N1 viruses except subclade 7.2 (Fig. 1A) and defined a conformational 65C6 epitope (1). In this study, we determined the 100F4 epitope and dissected the neutralization mechanism by these antibodies.To map the 100F4 epitope, a yeast display analysis was carried out similarly to the way we mapped the 65C6 epitope (1, 2). Figure 1B shows the 15 single amino acid mutations in H5 hemagglutinin (HA) that abolish the binding of antibody 100F4. Among these, the 7 residues at positions 68,112, 137, 143, 251, 254, and 255 were on the HA surface, while the rest were underneath the surface.To test whether these 7 surface mutations would affect neutralization by antibody 100F4, genes encoding 7 full-length H5 HA single mutants derived from H5N1 strain A/Beijing/01/03 subclade 7.1 were constructed and used to generate H5N1 pseudotypes. The resistance of H5N1 pseudotypes to antibody 100F4 was measured with the pseudotype-based neutralization assay (3). Compared to the wild-type subclade 7.1 H5N1 pseudotype, only H5N1 pseudotypes expressing H5 HA mutants with mutations at position 68 or 112 (72 or 116 according to H3 numbering) were dramatically resistant to antibody 100F4 (Fig. 1C and D). On the HA surface, these two resistant residues are adjacent to each other (Fig. 1E), but they are next to the Cb in H1 HA and site E in H3 HA (4-7) ( Fig. 1F and G). The 100F4 epitope does not overlap any known epitopes in the head region detected by human and mouse MAb ( Fig. 1H and I). Thus, the 100F4 epitope is a new conserved conformational epitope on the globular head and away from the receptor binding site (RBS). In contrast, the 65C6 epitope partially overlaps with Sa in H1 HA at residue 161 (K165 according to H3 numbering) and with site A in H3 HA at residues 118 and 121 (T122 and F125 according to H3 numbering) (4-7). In addition, the 65C6 epitope also partially overlaps epitopes detected by some human MAb, i.e., FLA5.10 at P118 (P122 according to H3 numbering), FLD21.140 at S121, Y164, and T167 (S125, Y168, and T171 according to H3 numbering) (8), AVFLuigG01 at P118, Y164, and T167 (P122, ...

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“…Broad domain mapping for independent panels of anti-influenza mAbs are described from yeast libraries displaying fulllength, precursor influenza HA (HA0) or the HA1 and HA2 subunits separately (113,114). After confirming binding and domain localization, sublibraries of mutant HA fragments were generated by error-prone PCR for fine epitope mapping.…”

Section: Anti-influenza Mab Discovery Involving Yeast Displaymentioning

confidence: 99%

Phage and Yeast Display

Sheehan

Marasco

2015

Microbiol Spectr

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Despite the availability of antimicrobial drugs, the continued development of microbial resistance-established through escape mutations and the emergence of resistant strains-limits their clinical utility. The discovery of novel, therapeutic, monoclonal antibodies (mAbs) offers viable clinical alternatives in the treatment and prophylaxis of infectious diseases. Human mAb-based therapies are typically nontoxic in patients and demonstrate high specificity for the intended microbial target. This specificity prevents negative impacts on the patient microbiome and avoids driving the resistance of nontarget species. The in vitro selection of human antibody fragment libraries displayed on phage or yeast surfaces represents a group of well-established technologies capable of generating human mAbs. The advantage of these forms of microbial display is the large repertoire of human antibody fragments present during a single selection campaign. Furthermore, the in vitro selection environments of microbial surface display allow for the rapid isolation of antibodies-and their encoding genes-against infectious pathogens and their toxins that are impractical within in vivo systems, such as murine hybridomas. This article focuses on the technologies of phage display and yeast display, as these strategies relate to the discovery of human mAbs for the treatment and vaccine development of infectious diseases.

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Fine epitope mapping of monoclonal antibodies against hemagglutinin of a highly pathogenic H5N1 influenza virus using yeast surface display

Cited by 31 publications

References 12 publications

A Unique and Conserved Neutralization Epitope in H5N1 Influenza Viruses Identified by an Antibody against the A/Goose/Guangdong/1/96 Hemagglutinin

A Unique and Conserved Neutralization Epitope in H5N1 Influenza Viruses Identified by an Antibody against the A/Goose/Guangdong/1/96 Hemagglutinin

Unraveling of a Neutralization Mechanism by Two Human Antibodies against Conserved Epitopes in the Globular Head of H5 Hemagglutinin

Phage and Yeast Display

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