Deep mutational scanning of whole SARS-CoV-2 spike in an inverted infection system

bioRxiv (Cold Spring Harbor Laboratory), Год журнала: 2023, Номер unknown

Опубликована: Июль 18, 2023

Summary In order to investigate SARS-CoV-2 mutations and their impact on immune evasion infectivity, we developed a Deep Mutational Scanning (DMS) platform utilizing an inverted infection assay measure spike expression, ACE2 affinity, viral infectivity in human cells. Surprisingly, our analysis reveals that protein rather than is the primary factor affecting correlated with evolution. Notably, within N-terminal domain (NTD), expression infectivity-enhancing are concentrated flexible loops. We also observed Omicron variants BA.1 BA.2 exhibit through receptor binding (RBD) mutations, although these reduce structural stability. Interestingly, NTD has evolved increase stability, compensating for RBD instability resulting heightened overall infectivity. Our findings, available SpikeScanDB, emphasize importance of levels compensatory both domains shaping variant

Язык: Английский

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Understanding epistatic networks in the B1 β-lactamases through coevolutionary statistical modeling and deep mutational scanning

bioRxiv (Cold Spring Harbor Laboratory), Год журнала: 2023, Номер unknown

Опубликована: Ноя. 28, 2023

Abstract Over the course of evolution, proteins families undergo sequence diversification via mutation accumulation, with extant homologs often sharing less than 25% identity. The resulting diversity presents a complex view sequence-structure-function relationships, as epistasis is prevalent, and deleterious mutations in one protein can be tolerated homologous sequences through networks intramolecular, compensatory interactions. Understanding these epistatic crucial for understanding predicting function, yet comprehensive analysis such across limited. In this study, we combine computational experimental approaches to examine class B1 metallo-β-lactamases, diverse family antibiotic-degrading enzymes. Using Direct Coupling Analysis, assess global coevolutionary signatures family. We also obtain detailed data from deep mutational scanning on two distant homologs, NDM-1 VIM-2. There good agreement between approaches, revealing both family-wide homolog specific patterns that associated 3D structure. However, interactions remain complex, strong evolutionarily entrenched residues are not easily compensated by changes nearby

Язык: Английский

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Functional and antigenic characterization of SARS-CoV-2 spike fusion peptide by deep mutational scanning

bioRxiv (Cold Spring Harbor Laboratory), Год журнала: 2023, Номер unknown

Опубликована: Ноя. 29, 2023

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

Язык: Английский

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Mammalian Antigen Display for Pandemic Countermeasures

Methods in molecular biology, Год журнала: 2024, Номер unknown, С. 191 - 216

Опубликована: Янв. 1, 2024

Язык: Английский

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Multiplex Functional Characterization of Protein Variant Libraries in Mammalian Cells with Single-Copy Genomic Integration and High-Throughput DNA Sequencing

Methods in molecular biology, Год журнала: 2024, Номер unknown, С. 135 - 152

Опубликована: Янв. 1, 2024

Язык: Английский

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