Generation of monoclonal antibodies specific of the postfusion conformation of the Pneumovirinae fusion (F) protein

• Published in: Journal of virological methods Vol. 224; pp. 1 - 8
• Main Authors: Rodríguez, Laura, Olmedillas, Eduardo, Mas, Vicente, Vázquez, Mónica, Cano, Olga, Terrón, María C., Luque, Daniel, Palomo, Concepción, Melero, José A.
• Format: Journal Article
• Language: English
• Published: Netherlands Elsevier B.V 01-11-2015
• Subjects: Animals; Antibodies, Monoclonal - isolation & purification; Antibodies, Viral - isolation & purification; Conformation specific antibodies; Female; Fusion proteins; Human metapneumovirus; Mice, Inbred BALB C; Paramyxovirus; Pneumovirinae; Pneumovirinae - immunology; Protein Conformation; Rabbits; Respiratory syncytial virus; Viral Fusion Proteins - chemistry; Viral Fusion Proteins - immunology; Pneumovirinae; Conformation specific antibodies; Fusion proteins
• ISSN: 0166-0934; 1879-0984
• DOI: 10.1016/j.jviromet.2015.08.002

•The Pneumoviriniae fusion (F) protein mediates virus-cell membrane fusion.•The F protein refolds from a prefusion to a postfusion conformation during fusion.•Methods are described to isolate MAbs specific of the postfusion conformation.•Similar methods may be applicable to other type I viral fusion proteins.
Paramyxovirus entry into cells requires fusion of the viral and cell membranes mediated by one of the major virus glycoproteins, the fusion (F) glycoprotein which transits from a metastable pre-fusion conformation to a highly stable post-fusion structure during the membrane fusion process. F protein refolding involves large conformational changes of the protein trimer. One of these changes results in assembly of two heptad repeat sequences (HRA and HRB) from each protomer into a six-helix bundle (6HB) motif. To assist in distinguishing pre- and post-fusion conformations of the Pneumovirinae F proteins, and as extension of previous work (Palomo et al., 2014), a general strategy was designed to obtain polyclonal and particularly monoclonal antibodies specific of the 6HB motif of the Pneumovirinae fusion protein. The antibodies reported here should assist in the characterization of the structural changes that the F protein of human metapneumovirus or respiratory syncytial virus experiences during the process of membrane fusion.