RESUMO
Many antibody-based antivirals, including broadly neutralizing antibodies (bnAbs) against various influenza virus strains, suffer from limited potency. A booster of the antiviral activity of an antibody is expected to facilitate development of antiviral therapeutics. In this study, a nanodisc (ND), a discoidal lipid bilayer encircled by membrane scaffold proteins, is engineered to provide virucidal properties to antibodies, thereby augmenting their antiviral activity. NDs carrying the Fc-binding peptide sequence form an antibody-ND complex (ANC), which can co-endocytose into cells infected with influenza virus. ANC efficiently inhibits endosome escape of viral RNA by dual complimentary mode of action. While the antibody moiety in an ANC inhibits hemagglutinin-mediated membrane fusion, its ND moiety destroys the viral envelope using free hemagglutinins that are not captured by antibodies. Providing virus-infected host cells with the ability to self-eliminate by the synergistic effect of ANC components dramatically amplifies the antiviral efficacy of a bnAb against influenza virus. When the efficacy of ANC is assessed in mouse models, administration of ANCs dramatically reduces morbidity and mortality compared to bnAb alone. This study is the first to demonstrate the novel nanoparticle ANC and its role in combating viral infections, suggesting that ANC is a versatile platform applicable to various viruses.
Assuntos
Anticorpos Antivirais , Envelope Viral , Animais , Anticorpos Antivirais/farmacologia , Antivirais/farmacologia , Anticorpos Amplamente Neutralizantes , Hemaglutininas , CamundongosRESUMO
Molecules interfering with lipid bilayer function exhibit strong antiviral activity against a broad range of enveloped viruses, with a lower risk of resistance development than that for viral protein-targeting drugs. Amphipathic peptides are rich sources of such membrane-interacting antivirals. Here, we report that influenza viruses were effectively inactivated by M2 AH, an amphipathic peptide derived from the M2 protein of the influenza virus. Although overall hydrophobicity (
Assuntos
Antivirais/farmacologia , Membrana Celular/efeitos dos fármacos , Vírus da Influenza A Subtipo H1N1/efeitos dos fármacos , Peptídeos/farmacologia , Proteínas da Matriz Viral/química , Sequência de Aminoácidos , Animais , Antivirais/síntese química , Membrana Celular/química , Membrana Celular/virologia , Cães , Interações Hidrofóbicas e Hidrofílicas , Vírus da Influenza A Subtipo H1N1/crescimento & desenvolvimento , Vírus da Influenza A Subtipo H1N1/ultraestrutura , Concentração Inibidora 50 , Bicamadas Lipídicas/química , Lipossomos/química , Células Madin Darby de Rim Canino , Peptídeos/síntese química , Relação Estrutura-Atividade , Carga Viral/efeitos dos fármacosRESUMO
2'-Fucosyllactose (2'-FL) is the most abundant milk oligosaccharide in human breast milk and it has several benefits for infant health. The quantification of 2'-FL in breast milk or in samples from other sources generally requires lengthy analyses. These methods cannot be used to simultaneously detect 2'-FL in numerous samples, which would be more time-efficient. In this study, two genes, namely α1,2-fucosidase from Xanthomonas manihotis and l-fucose dehydrogenase from Pseudomonas sp. no. 1143, were identified, cloned and overexpressed in E. coli. The recombinant enzymes were produced as 6â¯×â¯His-tagged proteins and were purified to homogeneity using Ni2+ affinity chromatography. The purified α1,2-fucosidase and l-fucose dehydrogenase are monomers with molecular masses of 63â¯kDa and 36â¯kDa, respectively. Both enzymes have sufficiently high activities in phosphate-buffered saline (pH 7.0) at 37⯰C, making it possible to develop a coupled enzyme reaction in a single buffer system for the quantitative determination of 2'-FL in a large number of samples simultaneously. This method can be used to quantify 2'-FL in infant formulas and in samples collected from different phases of the biotechnological production of this oligosaccharide. Furthermore, the method is applicable for the rapid screening of active variants during the development of microbial strains producing 2'-FL.
Assuntos
Ensaios Enzimáticos , Fórmulas Infantis/química , Leite Humano/química , Trissacarídeos/análise , Desidrogenases de Carboidrato/química , Humanos , Lactente , Recém-Nascido , Pseudomonas/metabolismo , Xanthomonas axonopodis/metabolismo , alfa-L-Fucosidase/químicaRESUMO
Membrane-disrupting agents that selectively target virus versus host membranes could potentially inhibit a broad-spectrum of enveloped viruses, but currently such antivirals are lacking. Here, we develop a nanodisc incorporated with a decoy virus receptor that inhibits virus infection. Mechanistically, nanodiscs carrying the viral receptor sialic acid bind to influenza virions and are co-endocytosed into host cells. At low pH in the endosome, the nanodiscs rupture the viral envelope, trapping viral RNAs inside the endolysosome for enzymatic decomposition. In contrast, liposomes containing a decoy receptor show weak antiviral activity due to the lack of membrane disruption. The nanodiscs inhibit influenza virus infection and reduce morbidity and mortality in a mouse model. Our results suggest a new class of antivirals applicable to other enveloped viruses that cause irreversible physical damage specifically to virus envelope by viruses' own fusion machine. In conclusion, the lipid nanostructure provides another dimension for antiviral activity of decoy molecules.