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1.
bioRxiv ; 2024 Aug 31.
Artículo en Inglés | MEDLINE | ID: mdl-39257766

RESUMEN

Antibody discovery is crucial for developing therapeutics and vaccines as well as understanding adaptive immunity. However, the lack of approaches to synthesize antibodies with defined sequences in a high-throughput manner represents a major bottleneck in antibody discovery. Here, we presented oPool+ display, which combines oligo pool synthesis and mRNA display to construct and characterize many natively paired antibodies in parallel. As a proof-of-concept, we applied oPool+ display to rapidly screen the binding activity of >300 natively paired influenza hemagglutinin (HA) antibodies against the conserved HA stem domain. Structural analysis of 16.ND.92, one of the identified HA stem antibodies, revealed a unique binding mode distinct from other known broadly neutralizing HA stem antibodies with convergent sequence features. Yet, despite such differences, 16.ND.92 remained broadly reactive and conferred in vivo protection. Overall, this study not only established an experimental platform that can be applied in both research and therapeutics to accelerate antibody discovery, but also provides molecular insights into antibody responses to the influenza HA stem, which is a major target for universal influenza vaccine development.

2.
Front Bioeng Biotechnol ; 12: 1428832, 2024.
Artículo en Inglés | MEDLINE | ID: mdl-39119275

RESUMEN

This paper will focus on analyzing the argument with appealing to nature against synthetic biology and provide a counter-argument against it through demonstrating the ambiguity of the concept of nature, denying the existence of a morally significant line between natural and non/unnatural, and disproving the allegations against synthetic biology raised by the argument appealing to nature. The paper consists of two parts following a brief introduction. The first part will describe the argument appealing to nature against synthetic biology, and identify the deficiencies of the argument per se, e.g., the ambiguity of the concept 'nature'; and the problems in the morally significant line between the natural and the non/unnatural. The second part will discuss the allegations to synthetic biology stemming from this argument, e.g., committing metaphysical and ethical mistakes, and doing possible harms to the environment.

3.
Immunity ; 2024 Aug 15.
Artículo en Inglés | MEDLINE | ID: mdl-39163866

RESUMEN

Despite decades of antibody research, it remains challenging to predict the specificity of an antibody solely based on its sequence. Two major obstacles are the lack of appropriate models and the inaccessibility of datasets for model training. In this study, we curated >5,000 influenza hemagglutinin (HA) antibodies by mining research publications and patents, which revealed many distinct sequence features between antibodies to HA head and stem domains. We then leveraged this dataset to develop a lightweight memory B cell language model (mBLM) for sequence-based antibody specificity prediction. Model explainability analysis showed that mBLM could identify key sequence features of HA stem antibodies. Additionally, by applying mBLM to HA antibodies with unknown epitopes, we discovered and experimentally validated many HA stem antibodies. Overall, this study not only advances our molecular understanding of the antibody response to the influenza virus but also provides a valuable resource for applying deep learning to antibody research.

4.
Nat Commun ; 15(1): 5175, 2024 Jun 18.
Artículo en Inglés | MEDLINE | ID: mdl-38890325

RESUMEN

The receptor-binding site of influenza A virus hemagglutinin partially overlaps with major antigenic sites and constantly evolves. In this study, we observe that mutations G186D and D190N in the hemagglutinin receptor-binding site have coevolved in two recent human H3N2 clades. X-ray crystallography results show that these mutations coordinately drive the evolution of the hemagglutinin receptor binding mode. Epistasis between G186D and D190N is further demonstrated by glycan binding and thermostability analyses. Immunization and neutralization experiments using mouse and human samples indicate that the evolution of receptor binding mode is accompanied by a change in antigenicity. Besides, combinatorial mutagenesis reveals that G186D and D190N, along with other natural mutations in recent H3N2 strains, alter the compatibility with a common egg-adaptive mutation in seasonal influenza vaccines. Overall, our findings elucidate the role of epistasis in shaping the recent evolution of human H3N2 hemagglutinin and substantiate the high evolvability of its receptor-binding mode.


Asunto(s)
Epistasis Genética , Evolución Molecular , Glicoproteínas Hemaglutininas del Virus de la Influenza , Subtipo H3N2 del Virus de la Influenza A , Gripe Humana , Humanos , Subtipo H3N2 del Virus de la Influenza A/genética , Subtipo H3N2 del Virus de la Influenza A/metabolismo , Glicoproteínas Hemaglutininas del Virus de la Influenza/genética , Glicoproteínas Hemaglutininas del Virus de la Influenza/química , Glicoproteínas Hemaglutininas del Virus de la Influenza/metabolismo , Animales , Ratones , Sitios de Unión , Gripe Humana/virología , Mutación , Cristalografía por Rayos X , Vacunas contra la Influenza , Unión Proteica , Receptores Virales/metabolismo , Receptores Virales/genética , Receptores Virales/química , Femenino
5.
Nat Commun ; 15(1): 4056, 2024 May 14.
Artículo en Inglés | MEDLINE | ID: mdl-38744813

RESUMEN

The fusion peptide of SARS-CoV-2 spike protein is functionally important for membrane fusion during virus entry and is part of a broadly neutralizing epitope. However, sequence determinants at the fusion peptide and its adjacent regions for pathogenicity and antigenicity remain elusive. In this study, we perform a series of deep mutational scanning (DMS) experiments on an S2 region spanning the fusion peptide of authentic SARS-CoV-2 in different cell lines and in the presence of broadly neutralizing antibodies. We identify mutations at residue 813 of the spike protein that reduced TMPRSS2-mediated entry with decreased virulence. In addition, we show that an F823Y mutation, present in bat betacoronavirus HKU9 spike protein, confers resistance to broadly neutralizing antibodies. Our findings provide mechanistic insights into SARS-CoV-2 pathogenicity and also highlight a potential challenge in developing broadly protective S2-based coronavirus vaccines.


Asunto(s)
Anticuerpos Neutralizantes , COVID-19 , Mutación , SARS-CoV-2 , Glicoproteína de la Espiga del Coronavirus , Internalización del Virus , Glicoproteína de la Espiga del Coronavirus/inmunología , Glicoproteína de la Espiga del Coronavirus/genética , Glicoproteína de la Espiga del Coronavirus/química , Glicoproteína de la Espiga del Coronavirus/metabolismo , Humanos , SARS-CoV-2/inmunología , SARS-CoV-2/genética , Anticuerpos Neutralizantes/inmunología , COVID-19/virología , COVID-19/inmunología , Animales , Anticuerpos Antivirales/inmunología , Serina Endopeptidasas/genética , Serina Endopeptidasas/inmunología , Serina Endopeptidasas/metabolismo , Chlorocebus aethiops , Células HEK293 , Células Vero , Epítopos/inmunología , Epítopos/genética , Línea Celular , Ratones
6.
Proc Natl Acad Sci U S A ; 121(15): e2317222121, 2024 Apr 09.
Artículo en Inglés | MEDLINE | ID: mdl-38557175

RESUMEN

Antigenic drift of SARS-CoV-2 is typically defined by mutations in the N-terminal domain and receptor binding domain of spike protein. In contrast, whether antigenic drift occurs in the S2 domain remains largely elusive. Here, we perform a deep mutational scanning experiment to identify S2 mutations that affect binding of SARS-CoV-2 spike to three S2 apex public antibodies. Our results indicate that spatially diverse mutations, including D950N and Q954H, which are observed in Delta and Omicron variants, respectively, weaken the binding of spike to these antibodies. Although S2 apex antibodies are known to be nonneutralizing, we show that they confer protection in vivo through Fc-mediated effector functions. Overall, this study indicates that the S2 domain of SARS-CoV-2 spike can undergo antigenic drift, which represents a potential challenge for the development of more universal coronavirus vaccines.


Asunto(s)
Deriva y Cambio Antigénico , COVID-19 , Humanos , SARS-CoV-2/genética , Anticuerpos , Glicoproteína de la Espiga del Coronavirus/genética , Anticuerpos Antivirales
7.
bioRxiv ; 2023 Nov 29.
Artículo en Inglés | MEDLINE | ID: mdl-38076875

RESUMEN

The fusion peptide of SARS-CoV-2 spike protein is functionally important for membrane fusion during virus entry and is part of a broadly neutralizing epitope. However, sequence determinants at the fusion peptide and its adjacent regions for pathogenicity and antigenicity remain elusive. In this study, we performed a series of deep mutational scanning (DMS) experiments on an S2 region spanning the fusion peptide of authentic SARS-CoV-2 in different cell lines and in the presence of broadly neutralizing antibodies. We identified mutations at residue 813 of the spike protein that reduced TMPRSS2-mediated entry with decreased virulence. In addition, we showed that an F823Y mutation, present in bat betacoronavirus HKU9 spike protein, confers resistance to broadly neutralizing antibodies. Our findings provide mechanistic insights into SARS-CoV-2 pathogenicity and also highlight a potential challenge in developing broadly protective S2-based coronavirus vaccines.

8.
Cell Rep ; 42(11): 113410, 2023 11 28.
Artículo en Inglés | MEDLINE | ID: mdl-37976161

RESUMEN

IGHV1-69 is frequently utilized by broadly neutralizing influenza antibodies to the hemagglutinin (HA) stem. These IGHV1-69 HA stem antibodies have diverse complementarity-determining region (CDR) H3 sequences. Besides, their light chains have minimal to no contact with the epitope. Consequently, sequence determinants that confer IGHV1-69 antibodies with HA stem specificity remain largely elusive. Using high-throughput experiments, this study reveals the importance of light-chain sequence for the IGHV1-69 HA stem antibody CR9114, which is the broadest influenza antibody known to date. Moreover, we demonstrate that the CDR H3 sequences from many other IGHV1-69 antibodies, including those to the HA stem, are incompatible with CR9114. Along with mutagenesis and structural analysis, our results indicate that light-chain and CDR H3 sequences coordinately determine the HA stem specificity of IGHV1-69 antibodies. Overall, this work provides molecular insights into broadly neutralizing antibody responses to influenza virus, which have important implications for universal influenza vaccine development.


Asunto(s)
Vacunas contra la Influenza , Gripe Humana , Humanos , Hemaglutininas , Anticuerpos ampliamente neutralizantes , Anticuerpos Neutralizantes , Glicoproteínas Hemaglutininas del Virus de la Influenza , Anticuerpos Antivirales , Regiones Determinantes de Complementariedad
9.
Immunity ; 56(11): 2621-2634.e6, 2023 Nov 14.
Artículo en Inglés | MEDLINE | ID: mdl-37967533

RESUMEN

There is growing appreciation for neuraminidase (NA) as an influenza vaccine target; however, its antigenicity remains poorly characterized. In this study, we isolated three broadly reactive N2 antibodies from the plasmablasts of a single vaccinee, including one that cross-reacts with NAs from seasonal H3N2 strains spanning five decades. Although these three antibodies have diverse germline usages, they recognize similar epitopes that are distant from the NA active site and instead involve the highly conserved underside of NA head domain. We also showed that all three antibodies confer prophylactic and therapeutic protection in vivo, due to both Fc effector functions and NA inhibition through steric hindrance. Additionally, the contribution of Fc effector functions to protection in vivo inversely correlates with viral growth inhibition activity in vitro. Overall, our findings advance the understanding of NA antibody response and provide important insights into the development of a broadly protective influenza vaccine.


Asunto(s)
Subtipo H1N1 del Virus de la Influenza A , Vacunas contra la Influenza , Gripe Humana , Infecciones por Orthomyxoviridae , Humanos , Gripe Humana/prevención & control , Neuraminidasa , Infecciones por Orthomyxoviridae/prevención & control , Subtipo H3N2 del Virus de la Influenza A , Epítopos , Anticuerpos Antivirales , Anticuerpos Monoclonales , Vacunación , Glicoproteínas Hemaglutininas del Virus de la Influenza
10.
Front Genet ; 14: 1261623, 2023.
Artículo en Inglés | MEDLINE | ID: mdl-37928244

RESUMEN

Background: Human biobanks are an essential resource for contemporary medical research, crucial in treating and preventing human diseases and improving health. Public trust in human biobanks is a vital social prerequisite for their continued operation and related research. Methods: Drawing on the "leap of faith" theory proposed by Georg Simmel and Guido Möllering, this paper first examines the relationship between public trust and human biobanks and the process through which such trust is established. Subsequently, based on the results of this analysis, targeted policy recommendations are put forward to consolidate or enhance public trust in human biobanks. Results: Public trust in human biobanks stems from certain "good reasons," through which uncertainty and vulnerability are "suspended" by faith, leading to a leap toward the "land of expectations." In this progress, the critical factors in building and enhancing public trust in human biobanks are the public's propensity to trust, the inherent trustworthiness of human biobanks, and the security and interactivity of the trust environment. Conclusion: Public trust in human biobanks cannot be determined by any universal formula, as it is influenced by many factors, including intangible elements such as faith that defy empirical understanding. Nonetheless, public trust in human biobanks can be enhanced through measures such as fostering the public's propensity to trust, enhancing the inherent trustworthiness of human biobanks, establishing structural safeguards for the trust environment through ethical norms, systems, and supervision, and promoting public participation.

12.
Acta bioeth ; 29(2)oct. 2023.
Artículo en Inglés | LILACS-Express | LILACS | ID: biblio-1519847

RESUMEN

Biometric technology has transformed human biological characteristics into a new form of privacy, and the misuse of this technology poses challenges to protecting this new privacy. This article initially defines biometric technology and biometric characteristics, further demonstrating why biometric characteristics belong to personal privacy and how biometric technology poses challenges to its protection. Through analysis, this article argues that the essence of these challenges is the conflicts between the ethical principle of privacy protection and the ethical principle of maximizing social benefits. In order to address these challenges, it is necessary first to weigh the fundamental ethical principles. The two basic principles of privacy protection and maximizing social benefits are not mutual antagonism but hierarchy, and this hierarchy should be based on the principle of practical feasibility. That is, applying biometric technology should first meet the principle of practical feasibility and, on this premise, realize the principle of maximizing social benefits based on not infringing on the principle of privacy protection.


La tecnología biométrica ha transformado las características biológicas humanas en una nueva forma de privacidad, y el uso indebido de esta tecnología plantea desafíos a su protección. En este artículo se define inicialmente la tecnología biométrica y las características biométricas; se demuestra además por qué las características biométricas pertenecen a la privacidad personal y cómo la tecnología biométrica plantea retos para su protección. Este artículo argumenta que la esencia de estos retos es el conflicto entre el principio ético de protección de la privacidad y el de maximización de los beneficios sociales. Para abordar estos retos es necesario sopesar primero los principios éticos fundamentales. Los dos principios básicos de protección de la privacidad y maximización de los beneficios sociales no son antagónicos, sino jerárquicos, y esta jerarquía debe basarse en el principio de viabilidad práctica. Es decir, la aplicación de la tecnología biométrica debe cumplir primero el principio de viabilidad práctica y, a partir de esta premisa, realizar el principio de maximización de los beneficios sociales sobre la base de no infringir el principio de protección de la intimidad.


A tecnologia biométrica transformou as características biológicas humanas em uma nova forma de privacidade, e o mal uso dessa tecnologia apresenta desafios para proteger essa nova privacidade. Esse artigo inicialmente define tecnologia biométrica e características biométricas, demonstrando posteriormente por que características biométricas pertencem à privacidade pessoal e como tecnologia biométrica coloca desafios à sua proteção. Através de análise, esse artigo discute que a essência desses desafios é o conflito entre o princípio ético da proteção da privacidade e o princípio ético de maximizar benefícios sociais. De forma a visar esses desafios é necessário primeiro ponderar os princípios éticos fundamentais. Os dois princípios básicos de proteção da privacidade e de maximizar benefícios sociais não são mutuamente antagônicos mas hierárquicos, e essa hierarquia deve ser baseada no princípio da viabilidade prática. Isso é, aplicar tecnologia biométrica deve primeiro atender ao princípio da viabilidade prática e, nessa premissa, compreender o princípio de maximizar benefícios sociais com base em não infringir o princípio de proteção da privacidade.

13.
Front Pharmacol ; 14: 1214590, 2023.
Artículo en Inglés | MEDLINE | ID: mdl-37701030

RESUMEN

As international academic exchanges and cooperation deepen, China has actively engaged in international biomedical research collaboration and achieved significant success. However, these accomplishments have been accompanied by ethical controversies and issues, with ethics dumping being a recurrently discussed focus among scholars. This paper reviews ethics dumping incidents in China's biomedical research field and analyzes the underlying causes to answer why China is often susceptible to ethics dumping. We argue that the primary reasons include weak ethical awareness among some researchers, an oversimplified research evaluation system, gaps in relevant ethics governance and oversight mechanisms, and limited capabilities of certain ethics committees. To address these issues, we propose five ethics governance recommendations: establishing refined ethics committees at various levels and types; advancing theoretical and practical research on science and technology ethics governance; strengthening legislation and regulation related to emerging science and technology; emphasizing self-regulation and capacity building of research institutions; and providing special protection and healthcare for victims of ethics dumping. The aim is to enhance China's research supervision system and prevent similar ethics dumping incidents from recurring.

14.
bioRxiv ; 2023 Sep 14.
Artículo en Inglés | MEDLINE | ID: mdl-37745338

RESUMEN

Despite decades of antibody research, it remains challenging to predict the specificity of an antibody solely based on its sequence. Two major obstacles are the lack of appropriate models and inaccessibility of datasets for model training. In this study, we curated a dataset of >5,000 influenza hemagglutinin (HA) antibodies by mining research publications and patents, which revealed many distinct sequence features between antibodies to HA head and stem domains. We then leveraged this dataset to develop a lightweight memory B cell language model (mBLM) for sequence-based antibody specificity prediction. Model explainability analysis showed that mBLM captured key sequence motifs of HA stem antibodies. Additionally, by applying mBLM to HA antibodies with unknown epitopes, we discovered and experimentally validated many HA stem antibodies. Overall, this study not only advances our molecular understanding of antibody response to influenza virus, but also provides an invaluable resource for applying deep learning to antibody research.

15.
bioRxiv ; 2023 Jul 06.
Artículo en Inglés | MEDLINE | ID: mdl-37461670

RESUMEN

IGHV1-69 is frequently utilized by broadly neutralizing influenza antibodies to the hemagglutinin (HA) stem. These IGHV1-69 HA stem antibodies have diverse complementarity-determining region (CDR) H3 sequences. Besides, their light chains have minimal to no contact with the epitope. Consequently, sequence determinants that confer IGHV1-69 antibodies with HA stem specificity remain largely elusive. Using high-throughput experiments, this study revealed the importance of light chain sequence for the IGHV1-69 HA stem antibody CR9114, which is the broadest influenza antibody known to date. Moreover, we demonstrated that the CDR H3 sequences from many other IGHV1-69 antibodies, including those to HA stem, were incompatible with CR9114. Along with mutagenesis and structural analysis, our results indicate that light chain and CDR H3 sequences coordinately determine the HA stem specificity of IGHV1-69 antibodies. Overall, this work provides molecular insights into broadly neutralizing antibody responses to influenza virus, which have important implications for universal influenza vaccine development.

16.
Nat Commun ; 14(1): 2003, 2023 04 10.
Artículo en Inglés | MEDLINE | ID: mdl-37037866

RESUMEN

Designing prefusion-stabilized SARS-CoV-2 spike is critical for the effectiveness of COVID-19 vaccines. All COVID-19 vaccines in the US encode spike with K986P/V987P mutations to stabilize its prefusion conformation. However, contemporary methods on engineering prefusion-stabilized spike immunogens involve tedious experimental work and heavily rely on structural information. Here, we establish a systematic and unbiased method of identifying mutations that concomitantly improve expression and stabilize the prefusion conformation of the SARS-CoV-2 spike. Our method integrates a fluorescence-based fusion assay, mammalian cell display technology, and deep mutational scanning. As a proof-of-concept, we apply this method to a region in the S2 domain that includes the first heptad repeat and central helix. Our results reveal that besides K986P and V987P, several mutations simultaneously improve expression and significantly lower the fusogenicity of the spike. As prefusion stabilization is a common challenge for viral immunogen design, this work will help accelerate vaccine development against different viruses.


Asunto(s)
COVID-19 , SARS-CoV-2 , Animales , Humanos , COVID-19/prevención & control , Vacunas contra la COVID-19 , Glicoproteína de la Espiga del Coronavirus , Mutación , Mamíferos/metabolismo
17.
mBio ; 14(3): e0008423, 2023 06 27.
Artículo en Inglés | MEDLINE | ID: mdl-37070986

RESUMEN

Immune imprinting is a driver known to shape the anti-hemagglutinin (HA) antibody landscape of individuals born within the same birth cohort. With the HA and neuraminidase (NA) proteins evolving at different rates under immune selection pressures, anti-HA and anti-NA antibody responses since childhood influenza virus infections have not been evaluated in parallel at the individual level. This is partly due to the limited knowledge of changes in NA antigenicity, as seasonal influenza vaccines have focused on generating neutralizing anti-HA antibodies against HA antigenic variants. Here, we systematically characterized the NA antigenic variants of seasonal A(H1N1) viruses from 1977 to 1991 and completed the antigenic profile of N1 NAs from 1977 to 2015. We identified that NA proteins of A/USSR/90/77, A/Singapore/06/86, and A/Texas/36/91 were antigenically distinct and mapped N386K as a key determinant of the NA antigenic change from A/USSR/90/77 to A/Singapore/06/86. With comprehensive panels of HA and NA antigenic variants of A(H1N1) and A(H1N1)pdm09 viruses, we determined hemagglutinin inhibition (HI) and neuraminidase inhibition (NI) antibodies from 130 subjects born between 1950 and 2015. Age-dependent imprinting was observed for both anti-HA and anti-NA antibodies, with the peak HI and NI titers predominantly detected from subjects at 4 to 12 years old during the year of initial virus isolation, except the age-independent anti-HA antibody response against A(H1N1)pdm09 viruses. More participants possessed antibodies that reacted to multiple antigenically distinct NA proteins than those with antibodies that reacted to multiple antigenically distinct HA proteins. Our results support the need to include NA proteins in seasonal influenza vaccine preparations. IMPORTANCE Seasonal influenza vaccines have aimed to generate neutralizing anti-HA antibodies for protection since licensure. More recently, anti-NA antibodies have been established as an additional correlate of protection. While HA and NA antigenic changes occurred discordantly, the anti-HA and anti-NA antibody profiles have rarely been analyzed in parallel at the individual level, due to the limited knowledge on NA antigenic changes. By characterizing NA antigenic changes of A(H1N1) viruses, we determined the anti-HA and anti-NA antibody landscape against antigenically distinct A(H1N1) and A(H1N1)pdm09 viruses using sera of 130 subjects born between 1950 and 2015. We observed age-dependent imprinting of both anti-HA and anti-NA antibodies against strains circulated during the first decade of life. A total of 67.7% (88/130) and 90% (117/130) of participants developed cross-reactive antibodies to multiple HA and NA antigens at titers ≥1:40. With slower NA antigenic changes and cross-reactive anti-NA antibody responses, including NA protein in influenza vaccine preparation may enhance vaccine efficacy.


Asunto(s)
Subtipo H1N1 del Virus de la Influenza A , Vacunas contra la Influenza , Gripe Humana , Humanos , Niño , Preescolar , Hemaglutininas , Formación de Anticuerpos , Neuraminidasa/genética , Anticuerpos Antivirales , Anticuerpos Neutralizantes , Glicoproteínas Hemaglutininas del Virus de la Influenza/genética
18.
Cell Rep ; 42(1): 111951, 2023 01 31.
Artículo en Inglés | MEDLINE | ID: mdl-36640354

RESUMEN

Influenza neuraminidase (NA) has received increasing attention as an effective vaccine target. However, its mutational tolerance is not well characterized. Here, the fitness effects of >6,000 mutations in human H3N2 NA are probed using deep mutational scanning. Our result shows that while its antigenic regions have high mutational tolerance, there are solvent-exposed regions with low mutational tolerance. We also find that protein stability is a major determinant of NA mutational fitness. The deep mutational scanning result correlates well with mutational fitness inferred from natural sequences using a protein language model, substantiating the relevance of our findings to the natural evolution of circulating strains. Additional analysis further suggests that human H3N2 NA is far from running out of mutations despite already evolving for >50 years. Overall, this study advances our understanding of the evolutionary potential of NA and the underlying biophysical constraints, which in turn provide insights into NA-based vaccine design.


Asunto(s)
Gripe Humana , Humanos , Gripe Humana/genética , Subtipo H3N2 del Virus de la Influenza A/genética , Neuraminidasa/genética , Neuraminidasa/metabolismo , Evolución Molecular , Mutación/genética
19.
Sci Adv ; 8(47): eadd7221, 2022 Nov 25.
Artículo en Inglés | MEDLINE | ID: mdl-36417523

RESUMEN

Increasing the expression level of the SARS-CoV-2 spike (S) protein has been critical for COVID-19 vaccine development. While previous efforts largely focused on engineering the receptor-binding domain (RBD) and the S2 subunit, the amino-terminal domain (NTD) has been long overlooked because of the limited understanding of its biophysical constraints. In this study, the effects of thousands of NTD single mutations on S protein expression were quantified by deep mutational scanning. Our results revealed that in terms of S protein expression, the mutational tolerability of NTD residues was inversely correlated with their proximity to the RBD and S2. We also identified NTD mutations at the interdomain interface that increased S protein expression without altering its antigenicity. Overall, this study not only advances the understanding of the biophysical constraints of the NTD but also provides invaluable insights into S-based immunogen design.

20.
Nat Commun ; 13(1): 6443, 2022 10 28.
Artículo en Inglés | MEDLINE | ID: mdl-36307418

RESUMEN

Neuraminidase (NA) of human influenza H3N2 virus has evolved rapidly and been accumulating mutations for more than half-century. However, biophysical constraints that govern the evolutionary trajectories of NA remain largely elusive. Here, we show that among 70 natural mutations that are present in the NA of a recent human H3N2 strain, >10% are deleterious for an ancestral strain. By mapping the permissive mutations using combinatorial mutagenesis and next-generation sequencing, an extensive epistatic network is revealed. Biophysical and structural analyses further demonstrate that certain epistatic interactions can be explained by non-additive stability effect, which in turn modulates membrane trafficking and enzymatic activity of NA. Additionally, our results suggest that other biophysical mechanisms also contribute to epistasis in NA evolution. Overall, these findings not only provide mechanistic insights into the evolution of human influenza NA and elucidate its sequence-structure-function relationship, but also have important implications for the development of next-generation influenza vaccines.


Asunto(s)
Vacunas contra la Influenza , Gripe Humana , Humanos , Neuraminidasa , Gripe Humana/epidemiología , Subtipo H3N2 del Virus de la Influenza A/genética , Prevalencia
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