title: Electrochemical detection of the oligomerization of PB1-F2 influenza A virus protein in infected cells.
abstract: PB1-F2 is a nonstructural accessory protein of Influenza A virus described to enhance the mortality and the morbidity of the virus in a host-dependent manner. In this work, an electrochemical biosensor based on an immunodetection system was developed to follow the oligomerization of PB1-F2 during the viral cycle. The immunosensor was based on conductive polypyrrole modified with ferrocenyl groups as a redox marker for enhancing signal detection. Antibodies specific for monomeric or oligomeric PB1-F2 forms were immobilized on polypyrrole matrix via biotin/streptavidin layer. We demonstrated that this electrochemical biosensor sensitively detects PB1-F2 in both conformational forms. The linear range extends from 5 nM to 1.5 μM and from 5 nM to 0.5 μM for monomeric and oligomeric PB1-F2, respectively. The calculated limit of detection was 0.42 nM for monomeric PB1-F2 and 16 nM for oligomers. The biosensor platform allows the detection and quantification of PB1-F2 in lysates of infected cells during viral cycle. We show that at early stages of viral cycle, PB1-F2 is mainly monomeric but switched to amyloid-like structures at a later stage of infection. The quantification of two protein structural forms points out that PB1-F2 expression profiles and kinetics of oligomerization are cell-type-dependent.
authors: CNRS UMR-8182, Institut de Chimie Moléculaire et de Matériaux d'Orsay, Equipe de Chimie Bioorganique et Bioinorganique, Bâtiment 420, 91405 Orsay, France.
authors: Anna
authors: A
authors: Miodek
authors: Jasmina
authors: J
authors: Vidic
authors: Helene
authors: H
authors: Sauriat-Dorizon
authors: Charles-Adrien
authors: CA
authors: Richard
authors: Ronan
authors: R
authors: Le Goffic
authors: Hafsa
authors: H
authors: Korri-Youssoufi
authors: Christophe
authors: C
authors: Chevalier
mesh_terms: D000911:Antibodies, Monoclonal / Q000276:immunology; D052160:Benzothiazoles; D015374:Biosensing Techniques; D045744:Cell Line, Tumor; D055664:Electrochemical Techniques; D006801:Humans; D009980:Influenza A virus / Q000502:physiology*; D018625:Microscopy, Atomic Force; D055503:Protein Multimerization; D011994:Recombinant Proteins / Q000032:analysis / Q000096:biosynthesis / Q000235:genetics; D013844:Thiazoles / Q000737:chemistry; D014764:Viral Proteins / Q000032:analysis* / Q000235:genetics / Q000378:metabolism
chemical_list: D000911:Antibodies, Monoclonal; D052160:Benzothiazoles; C442816:PB1-F2 protein, Influenza A virus; D011994:Recombinant Proteins; D013844:Thiazoles; D014764:Viral Proteins; C009462:thioflavin T